1,3,4-thiadiazole compounds and their use in treating cancer

ABSTRACT

A compound of Formula (I):or a pharmaceutically acceptable salt thereof, where: Q can be 1,2,4-triazin-3-yl, pyridazin-3-yl, 6-methylpyridazin-3-yl, or 6-fluoropyridazin-3-yl; R1 can be hydrogen, methoxy, trifluoromethoxy, oxetan-3-yl, 3-fluoroazetidin-1-yl, 3-methoxyazetidin-1-yl, or 3,3-difluoroazetidin-1-yl; R2 can be hydrogen or fluoro; R3 can be hydrogen or methoxy; and R4 can be methoxy, ethoxy, or methoxymethyl; provided that when R1 is hydrogen, methoxy or trifluoromethoxy, then R3 is not hydrogen, and/or R4 is methoxymethyl. The compound of formula (I) can inhibit glutaminase, e.g., GLS1.

TECHNICAL FIELD

The specification generally relates to substituted 1,3,4-thiadiazolecompounds and pharmaceutically acceptable salts thereof. These compoundsact on the glutaminase 1 enzyme (“GLS1”), and the specificationtherefore also relates to the use of such compounds and salts thereof totreat or prevent GLS1-mediated disease, including cancer. Thespecification further relates to pharmaceutical compositions comprisingsuch compounds and salts; kits comprising such compounds and salts;methods of manufacture of such compounds and salts; intermediates usefulin the manufacture of such compounds and salts; and to methods oftreating GLS1 mediated disease, including cancer, using such compoundsand salts.

BACKGROUND

Glutamine is the most abundant plasma amino acid and is involved in manygrowth promoting pathways. In particular, glutamine is involved inoxidation in the TCA cycle and in maintaining cell redox equilibrium,and also provides nitrogen for nucleotide and amino acid synthesis (Curiet al., Front. Biosci. 2007, 12, 344-57; DeBerardinis and Cheng,Oncogene 2010, 313-324, each of which is incorporated by reference inits entirety). Many cancer cells rely on glutamine metabolism as aconsequence of metabolic changes in the cell, including the Warburgeffect where glycolytic pyruvate is converted to lactic acid rather thanbeing used to create acetyl CoA (Koppenol et al., Nature Reviews 2011,11, 325-337, which is incorporated by reference in its entirety). As aconsequence of this reliance on glutamine metabolism, such cancer cellsare sensitive to changes in exogenous glutamine levels. Furthermore,existing evidence suggests that glutaminolysis plays a key role incertain cancer types (Hensley et al., J. Clin. Invest. 2013, 123,3678-3684, which is incorporated by reference in its entirety), and isassociated with known oncogenic drivers such as Myc (Dang, Cancer Res.2010, 70, 859-863, which is incorporated by reference in its entirety).

The first step of glutamine catabolism to glutamate is catalysed byglutaminase, which exists as two isoforms, GLS1 and GLS2, originallyidentified as being expressed in the kidney and liver, respectively.Kidney glutaminase (GLS1) is known to be more ubiquitously expressedthan liver glutaminase (GLS2), and has 2 splice variants, KGA and theshorter GAC isoform, both of which are located in the mitochondria.(Elgadi et al., Physiol. Genomics 1999, 1, 51-62; Cassago et al., Proc.Natl. Acad. Sci. 2012, 109, 1092-1097, each of which is incorporated byreference in its entirety). GLS1 expression is associated with tumourgrowth and malignancy in a number of disease types (Wang et al., CancerCell 2010, 18, 207-219; van der Heuval et al., Cancer Bio. Ther. 2012,13, 1185-1194, each of which is incorporated by reference in itsentirety). Inhibitors of GLS1 are therefore expected to be useful in thetreatment of cancer, as monotherapy or in combination with otheranti-cancer agents.

SUMMARY

In one aspect, a compound of Formula (I):

or a pharmaceutically acceptable salt thereof, where:

Q can be 1,2,4-triazin-3-yl, pyridazin-3-yl, 6-methylpyridazin-3-yl, or6-fluoropyridazin-3-yl;

R¹ can be hydrogen, methoxy, trifluoromethoxy, oxetan-3-yl,3-fluoroazetidin-1-yl, 3-methoxyazetidin-1-yl, or3,3-difluoroazetidin-1-yl;

R² can be hydrogen or fluoro;

R³ can be hydrogen or methoxy; and

R⁴ can be methoxy, ethoxy, or methoxymethyl;

provided that when R¹ is hydrogen, methoxy or trifluoromethoxy, then R³is not hydrogen, and/or R⁴ is methoxymethyl.

In another aspect, a compound, or a pharmaceutically acceptable saltthereof, selected from the group consisting of:

-   (2S)-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-[3-(trifluoromethoxy)phenyl]acetamide;-   (2R)-2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;    and-   (2S)-[3-(difluoromethoxy)phenyl]-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide.

In another aspect, a pharmaceutical composition includes a compound asdescribed above, or a pharmaceutically acceptable salt thereof, and atleast one pharmaceutically acceptable diluent or carrier.

In another aspect, a compound as described above, or a pharmaceuticallyacceptable salt thereof, for use in therapy.

In another aspect, a compound as described above, or a pharmaceuticallyacceptable salt thereof, for use in the treatment of cancer.

In another aspect, use of a compound as described above, or apharmaceutically acceptable salt thereof, for the manufacture of amedicament for the treatment of cancer.

In another aspect, a method for treating cancer in a warm blooded animalin need of such treatment, includes administering to the warm-bloodedanimal a therapeutically effective amount of a compound as describedabove, or a pharmaceutically acceptable salt thereof.

Other aspects will be apparent from the specification and the claims.

DETAILED DESCRIPTION

Many embodiments are detailed throughout the specification and will beapparent to a reader skilled in the art. The invention is not to beinterpreted as being limited to any particular embodiment(s) thereof.

A compound of Formula (I) is provided:

or a pharmaceutically acceptable salt thereof, where:

Q can be 1,2,4-triazin-3-yl, pyridazin-3-yl, 6-methylpyridazin-3-yl, or6-fluoropyridazin-3-yl;

R¹ can be hydrogen, methoxy, trifluoromethoxy, oxetan-3-yl,3-fluoroazetidin-1-yl; 3-methoxyazetidin-1-yl, or3,3-difluoroazetidin-1-yl;

R² can be hydrogen or fluoro;

R³ can be hydrogen or methoxy; and

R⁴ can be methoxy, ethoxy, methoxymethyl, or ethyoxymethyl;

provided that when R¹ is hydrogen, methoxy or trifluoromethoxy, then R³is not hydrogen, and/or R⁴ is methoxymethyl.

1,2,4-triazin-3-yl, pyridazin-3-yl, 6-methylpyridazin-3-yl, and6-fluoropyridazin-3-yl rings have the following structures:

Oxetan-3-yl, 3-fluoroazetidin-1-yl; 3-methoxyazetidin-1-yl, and3,3-difluoroazetidin-1-yl rings have the following structures:

In some embodiments, the compound of Formula (I) has the followingFormula (Ia):

wherein Q, R¹, R², R³, and R⁴ as defined as above.

A compound, or a pharmaceutically acceptable salt thereof, selected fromthe group consisting of:

-   (2S)-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-[3-(trifluoromethoxy)phenyl]acetamide;-   (2R)-2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;    and-   (2S)-[3-(difluoromethoxy)phenyl]-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;

is also provided.

The term “pharmaceutically acceptable” is used to specify that an object(for example a salt, dosage form, diluent or carrier) is suitable foruse in patients. An example list of pharmaceutically acceptable saltscan be found in the Handbook of Pharmaceutical Salts: Properties,Selection and Use, P. H. Stahl and C. G. Wermuth, editors,Weinheim/Ztrich: Wiley-VCH/VHCA, 2002, which is incorporated byreference in its entirety. A suitable pharmaceutically acceptable saltof a compound of Formula (I) is, for example, an acid-addition salt. Anacid addition salt of a compound of Formula (I) may be formed bybringing the compound into contact with a suitable inorganic or organicacid under conditions known to the skilled person. An acid addition saltmay be formed using, for example, an inorganic acid such as hydrochloricacid, hydrobromic acid, sulphuric acid, and phosphoric acid. An acidaddition salt may also be formed using, for example, an organic acidsuch as trifluoroacetic acid, methanesulfonic acid, or benzenesulfonicacid.

Therefore, in one embodiment there is provided a compound of Formula (I)or a pharmaceutically acceptable salt thereof, where thepharmaceutically acceptable salt is a hydrochloric acid, hydrobromicacid, sulphuric acid, phosphoric acid, trifluoroacetic acid,methanesulfonic acid, or benzenesulfonic acid salt.

In one embodiment there is provided a compound of Formula (I) or apharmaceutically acceptable salt thereof, where the pharmaceuticallyacceptable salt is a hydrochloric acid or hydrobromic acid salt.

A further suitable pharmaceutically acceptable salt of a compound ofFormula (I) is a base-addition salt. A base addition salt of a compoundof Formula (I) may be formed by bringing the compound into contact witha suitable inorganic or organic base under conditions known to theskilled person. A base addition salt may for example be formed using,for example, an inorganic base such as an alkali metal hydroxide (suchas sodium, potassium, or lithium hydroxide) or an alkaline earth metalhydroxide (such as calcium hydroxide or magnesium hydroxide). A baseaddition salt may also be formed using, for example, an organic basesuch as methylamine, dimethylamine, trimethylamine, piperidine,morpholine, or tris-(2-hydroxyethyl)amine.

Therefore, in one embodiment there is provided a compound of Formula (I)or a pharmaceutically acceptable salt thereof, where thepharmaceutically acceptable salt is a sodium hydroxide, potassiumhydroxide, lithium hydroxide, calcium hydroxide, magnesium hydroxide,methylamine, dimethylamine, trimethylamine, piperidine, morpholine, ortris-(2-hydroxyethyl)amine salt.

In one embodiment there is provided a compound of Formula (I) or apharmaceutically acceptable salt thereof, where the pharmaceuticallyacceptable salt is a hydrochloric acid, hydrobromic acid, sulphuricacid, phosphoric acid, trifluoroacetic acid, methanesulfonic acid,benzenesulfonic acid, sodium hydroxide, potassium hydroxide, lithiumhydroxide, calcium hydroxide, magnesium hydroxide, methylamine,dimethylamine, trimethylamine, piperidine, morpholine, ortris-(2-hydroxyethyl)amine salt.

A further embodiment provides any of the embodiments defined herein (forexample the embodiment of claim 1) with the proviso that one or morespecific Examples (for instance one, two or three specific Examples, oralternatively one specific Example) selected from the group consistingof Examples 1(a), 1(b), 2(a), 2(b), 3(a), 3(b), 4(a), 4(b), 5(a), 5(b),6(a), 6(b), 7(a), 7(b), 8(a), 8(b), 9(a), 9(b), 10(a), 10(b), 11(a),11(b), 12(a), 12(b), 13(a), 13(b), 14(a), 14(b), 15(a), 15(b), 16(a),16(b), 17(a), 17(b), 18(a), 18(b), 19(a), 19(b), 20(a), 20(b), 21(a),21(b), 22(a), 22(b), 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35,36, 37, 38, 39, 40, 41, 42, 43, 44, 45(a), 45(b), 46(a), 46(b), 47(a),47(b), 48(a), 48(b), 49(a), 49(b), 50(a), 50(b), 51(a), 51(b), 52(a),52(b), 53(a), 53(b), 54(a), and 54(b) is individually disclaimed.

Some values of variable groups in Formula (I) are as follows. Suchvalues may be used in combination with any of the definitions, claims(for example claim 1), or embodiments defined herein to provide furtherembodiments.

R¹ can be methoxy or trifluoromethoxy.

R¹ can be methoxy and R³ can be methoxy.

R¹ can be oxetan-3-yl, 3-fluoroazetidin-1-yl, 3-methoxyazetidin-1-yl, or3,3-difluoroazetidin-1-yl.

R¹ can be oxetan-3-yl.

R¹ can be 3-fluoroazetidin-1-yl.

R¹ can be 3-methoxyazetidin-1-yl.

R¹ can be 3,3-difluoroazetidin-1-yl.

R² can be H.

R² can be fluoro.

R³ can be methoxy.

R⁴ can be methoxy or ethoxy.

R⁴ can be methoxy.

R⁴ can be methoxymethyl.

Q can be 1,2,4-triazin-3-yl or pyridazin-3-yl.

Q can be 1,2,4-triazin-3-yl.

Q can be pyridazin-3-yl.

Q can be 6-methylpyridazin-3-yl or 6-fluoropyridazin-3-yl.

Q can be 6-methylpyridazin-3-yl.

Q can be 6-fluoropyridazin-3-yl.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, where:

Q is 1,2,4-triazin-3-yl, pyridazin-3-yl, 6-methylpyridazin-3-yl, or6-fluoropyridazin-3-yl;

R¹ is methoxy, trifluoromethoxy, oxetan-3-yl, 3-fluoroazetidin-1-yl,3-methoxyazetidin-1-yl, or 3,3-difluoroazetidin-1-yl;

R² is hydrogen or fluoro;

R³ is hydrogen or methoxy; and

R⁴ is methoxy, ethoxy, or methoxymethyl;

provided that when R¹ is methoxy or trifluoromethoxy, then R³ is nothydrogen, and/or R⁴ is methoxymethyl.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, where:

Q is pyridazin-3-yl; and

R¹ is methoxy or trifluoromethoxy.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, where:

Q is pyridazin-3-yl; and

R¹ is oxetan-3-yl, 3-fluoroazetidin-1-yl, 3-methoxyazetidin-1-yl, or3,3-difluoroazetidin-1-yl.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, where:

Q is pyridazin-3-yl;

R¹ is methoxy; and

R³ is methoxy.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, where:

Q is 6-methylpyridazin-3-yl or 6-fluoropyridazin-3-yl; and

R¹ is 3-fluoroazetidin-1-yl, 3-methoxyazetidin-1-yl, or3,3-difluoroazetidin-1-yl.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, where the compound is selectedfrom the group consisting of:

-   (2S)-2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-(3,5-dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-(3,5-dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2R)-3-methoxy-2-(3-methoxyphenyl)-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]propanamide;-   (2S)-2-(3,5-dimethoxyphenyl)-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-[4-fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-N-[5-[(3R)-3-(pyridazin-3-ylamino)pyrrolidin-1-yl]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)—N-[5-[[(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetamide;-   (2S)-2-[3-(3-fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-methoxy-2-[3-methoxy-5-(trifluoromethoxy)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2R)-2-methoxy-2-[3-methoxy-5-(trifluoromethoxy)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-(4-fluorophenyl)-2-methoxy-N-[5-[[(3R)-1-(5-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-[4-fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;    and-   (2S)-2-methoxy-2-[3-(oxetan-3-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, where the compound is selectedfrom the group consisting of:

-   (2S)-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-[3-(trifluoromethoxy)phenyl]acetamide;-   (2R)-2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;-   (2S)-2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide;    and-   (2S)-[3-(difluoromethoxy)phenyl]-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide.

Compounds and salts described in this specification may exist insolvated forms and unsolvated forms. For example, a solvated form may bea hydrated form, such as a hemi-hydrate, a mono-hydrate, a di-hydrate, atri-hydrate or an alternative quantity thereof. The present inventionencompasses all such solvated and unsolvated forms of compounds ofFormula (I).

Atoms of the compounds and salts described in this specification mayexist in different isotopic forms. The present invention encompasses allisotopic forms of compounds of Formula (I) including an ¹¹C or ¹³Ccarbon and ¹H, ²H (deuterium) or ³H (tritium)hydrogen.

Compounds and salts described in this specification may exist as amixture of tautomers. “Tautomers” are structural isomers that exist inequilibrium resulting from the migration of a hydrogen atom. The presentinvention includes all tautomers of compounds of Formula (I).

Compounds of Formula (I) can be prepared in different diastereomericforms. The present invention includes all diastereomeric forms of thecompounds of Formula (I).

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, which is a single diastereomerbeing in an diastereomeric excess (% de) of >95%, >98% or >99%. In oneembodiment, the single diastereomer is present in diastereomeric excess(% de) of >99%.

Compounds believed to inhibit GLS1, i.e., the compounds of Formula (I),and pharmaceutically acceptable salts thereof are expected to be usefulin therapy, for example in the treatment of diseases or medicalconditions mediated at least in part by GLS1, including cancer.

Where “cancer” is mentioned, this includes both non-metastatic cancerand also metastatic cancer, such that treating cancer involves treatmentof both primary tumours and also tumour metastases.

In one embodiment the cancer is metastatic cancer.

In one embodiment the cancer is non-metastatic cancer.

“GLS1 inhibitory activity” refers to a decrease in the activity of GLS1as a direct or indirect response to the presence of a compound ofFormula (I), or pharmaceutically acceptable salt thereof, relative tothe activity of GLS1 in the absence of compound of Formula (I), orpharmaceutically acceptable salt thereof. Such a decrease in activitymay be due to the direct interaction of the compound of Formula (I), orpharmaceutically acceptable salt thereof with GLS1, or due to theinteraction of the compound of Formula (I), or pharmaceuticallyacceptable salt thereof with one or more other factors that in turnaffect GLS1 activity. For example, the compound of Formula (I), orpharmaceutically acceptable salt thereof, may decrease GLS1 by directlybinding to GLS1; by causing (directly or indirectly) another factor todecrease GLS1 activity; or by (directly or indirectly) decreasing theamount of GLS1 present in the cell or organism.

The term “therapy” is intended to have its normal meaning of treating adisease or correcting or compensating for the underlying pathology. Theterm “therapy” also includes “prophylaxis” unless there are specificindications to the contrary. The terms “therapeutic” and“therapeutically” should be interpreted in a corresponding manner.

The term “therapeutically effective amount” refers to an amount of acompound of Formula (I) as described in any of the embodiments hereinwhich is effective to provide therapy in a subject. In the case ofcancer, the therapeutically effective amount may cause any of thechanges observable or measurable in a subject as described in thedefinition of “therapy”, “treatment” and “prophylaxis” above. Forexample, the effective amount can reduce the number of cancer or tumorcells; reduce the overall tumor size; inhibit or stop tumor cellinfiltration into peripheral organs including, for example, the softtissue and bone; inhibit and stop tumor metastasis; inhibit and stoptumor growth; relieve to some extent one or more of the symptomsassociated with the cancer; reduce morbidity and mortality; improvequality of life; or a combination of such effects. An effective amountmay be an amount sufficient to decrease the symptoms of a diseaseresponsive to inhibition of GLS1 activity. For cancer therapy, efficacyin-vivo can, for example, be measured by assessing the duration ofsurvival, time to disease progression (TTP), the response rates (RR),duration of response, and/or quality of life. As recognized by thoseskilled in the art, effective amounts may vary depending on route ofadministration, excipient usage, and co-usage with other agents. Forexample, where a combination therapy is used, the amount of the compoundof Formula (I) or pharmaceutically acceptable salt described in thisspecification and the amount of the other pharmaceutically activeagent(s) are, when combined, jointly effective to treat a targeteddisorder in the animal patient. In this context, the combined amountsare in a “therapeutically effective amount” if they are, when combined,sufficient to decrease the symptoms of a disease responsive toinhibition of GLS1 activity as described above. Typically, such amountsmay be determined by one skilled in the art by, for example, startingwith the dosage range described in this specification for the compoundof Formula (I) or pharmaceutically acceptable salt thereof and anapproved or otherwise published dosage range(s) of the otherpharmaceutically active compound(s).

The term “prophylaxis” is intended to have its normal meaning andincludes primary prophylaxis to prevent the development of the diseaseand secondary prophylaxis whereby the disease has already developed andthe patient is temporarily or permanently protected against exacerbationor worsening of the disease.

The term “treatment” is used synonymously with “therapy”. Similarly theterm “treat” can be regarded as applying therapy where “therapy” is asdefined herein.

In one embodiment there is provided a pharmaceutical compositionincluding the compound of Formula (I), or a pharmaceutically acceptablesalt thereof, and at least one pharmaceutically acceptable diluent orcarrier. In one embodiment, the pharmaceutical composition includes acompound of Formula (I) as a free base. In another embodiment, thepharmaceutical composition includes a a pharmaceutically acceptable saltof a compound of Formula (I).

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, for use in therapy.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, for use in the treatment ofcancer.

In one embodiment there is provided the use of the compound of Formula(I), or a pharmaceutically acceptable salt thereof, for the manufactureof a medicament for the treatment of cancer.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, for use in the treatment of adisease mediated by GLS1. In one embodiment, the disease mediated byGLS1 is cancer. In some embodiments, the cancer can be breast cancer(for example triple negative breast cancer), lung cancer (for examplenon-small cell lung cancer), pancreatic cancer, renal cancer, orhepatocellular cancer.

“Triple negative breast cancer” is any breast cancer that does notexpress, or underexpresses, the genes for the estrogen receptor,progesterone receptor and Her2/neu.

In one embodiment there is provided the use of the compound of Formula(I), or a pharmaceutically acceptable salt thereof, for the manufactureof a medicament for the treatment of a disease mediated by GLS1. In oneembodiment, the disease mediated by GLS1 is cancer. In some embodiments,the cancer can be breast cancer (for example triple negative breastcancer), lung cancer (for example non-small cell lung cancer),pancreatic cancer, renal cancer, or hepatocellular cancer.

In one embodiment there is provided the use of the compound of Formula(I), or a pharmaceutically acceptable salt thereof, for the manufactureof a medicament for the treatment of cancer.

In one embodiment there is provided a method of inhibiting GLS1 whichincludes administering a compound of Formula (I).

In one embodiment there is provided a method for treating a disease inwhich inhibition of GLS1 is beneficial in a warm-blooded animal in needof such treatment, which includes administering to the warm-bloodedanimal a therapeutically effective amount of a compound of Formula (I),or a pharmaceutically acceptable salt thereof.

“Warm-blooded animals” include, for example, humans.

In one embodiment there is provided a method for treating cancer in awarm-blooded animal in need of such treatment, which includesadministering to the warm-blooded animal a therapeutically effectiveamount of a compound of Formula (I), or a pharmaceutically acceptablesalt thereof. In some embodiments, the cancer can be breast cancer (forexample triple negative breast cancer), lung cancer (for examplenon-small cell lung cancer), pancreatic cancer, renal cancer, orhepatocellular cancer.

The treatment for cancer described in this specification may be appliedas a sole therapy, or may involve, in addition to administration of thecompound of Formula (I), conventional surgery, radiotherapy, orchemotherapy; or a combination of such additional therapies. Suchconventional surgery, radiotherapy, or chemotherapy may be administeredsimultaneously, sequentially, or separately to treatment with thecompound of Formula (I).

Therefore, in one embodiment there is provided a compound of Formula(I), or a pharmaceutically acceptable salt thereof, and at least oneadditional anti-tumour substance, for use in the treatment of cancer.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, and at least one additionalanti-tumour substance for use in the simultaneous, separate orsequential treatment of cancer.

In one embodiment there is provided a compound of Formula (I), or apharmaceutically acceptable salt thereof, for use in the treatment ofcancer, where the compound of Formula (I) is administeredsimultaneously, separately, or sequentially with at least one additionalanti-tumour substance.

In one embodiment there is provided a method of treating cancer in awarm-blooded animal who is in need of such treatment, which includesadministering to the warm-blooded animal a compound of Formula (I), or apharmaceutically acceptable salt thereof and at least one additionalanti-tumour substance, wherein the amounts of the compound of Formula(I), or a pharmaceutically acceptable salt thereof, and the additionalanti-tumour substance are jointly effective in producing an anti-cancereffect.

In one embodiment there is provided a method of treating cancer in awarm-blooded animal who is in need of such treatment, which includesadministering to the warm-blooded animal a compound of Formula (I), or apharmaceutically acceptable salt thereof, and simultaneously, separatelyor sequentially administering at least one additional anti-tumoursubstance to the warm-blooded animal, wherein the amounts of thecompound of Formula (I), or pharmaceutically acceptable salt thereof,and the additional anti-tumour substance are jointly effective inproducing an anti-cancer effect.

In any embodiment the additional anti-tumour substance is a taxane. Inone embodiment the taxane is paclitaxel. In one embodiment the taxane isdocetaxel.

In any embodiment the additional anti-tumour substance is a platinumtherapy. In one embodiment the platinum therapy is cisplatin,oxaliplatin, or carboplatin.

According to a further embodiment there is provided a kit comprising:

-   -   a) A compound of Formula (I), or a pharmaceutically acceptable        salt thereof, in a first unit dosage form;    -   b) A second anti-tumour substance in a second unit dosage form;    -   c) A container for containing the first and second unit dosage        forms; and, optionally,    -   d) Instructions for use.

The compounds of Formula (I), and pharmaceutically acceptable saltsthereof, may be administered as pharmaceutical compositions, comprisingone or more pharmaceutically acceptable diluents or carriers.Accordingly, in one embodiment there is provided a pharmaceuticalcomposition comprising a compound of Formula (I), or a pharmaceuticallyacceptable salt thereof, and at least one pharmaceutically acceptablediluent or carrier.

The compositions may be in a form suitable for oral use (for example astablets, lozenges, hard or soft capsules, aqueous or oily suspensions,emulsions, dispersible powders or granules, syrups or elixirs), fortopical use (for example as creams, ointments, gels, or aqueous or oilysolutions or suspensions), for administration by inhalation (for exampleas a finely divided powder or a liquid aerosol), for administration byinsufflation (for example as a finely divided powder) or for parenteraladministration (for example as a sterile aqueous or oily solution forintravenous, subcutaneous, intramuscular dosing), or as a suppository.The compositions may be obtained by conventional procedures usingconventional pharmaceutical excipients. Thus, compositions intended fororal use may contain, for example, one or more coloring, sweetening,flavoring, and/or preservative agents.

In one embodiment there is provided a pharmaceutical compositioncomprising a compound of Formula (I), or a pharmaceutically acceptablesalt thereof, and at least one pharmaceutically acceptable diluent orcarrier, for use in therapy.

In one embodiment there is provided a pharmaceutical compositioncomprising a compound of Formula (I), or a pharmaceutically acceptablesalt thereof, and at least one pharmaceutically acceptable diluent orcarrier, for use in the treatment of cancer. In some embodiments thecancer can be breast cancer (for example triple negative breast cancer),lung cancer (for example non-small cell lung cancer), pancreatic cancer,renal cancer, or hepatocellular cancer.

The compound of Formula (I) will normally be administered to awarm-blooded animal at a unit dose within the range 5-5000 mg/m² bodyarea of the animal, i.e., approximately 0.1-100 mg/kg, and this normallyprovides a therapeutically-effective dose. A unit dose form such as atablet or capsule will usually contain, for example 1-250 mg of activeingredient. The daily dose will necessarily be varied depending upon thehost treated, the particular route of administration, any therapiesbeing co-administered, and the severity of the illness being treated.Accordingly the practitioner who is treating any particular patient maydetermine the optimum dosage.

EXAMPLES

The various embodiments are illustrated by the following Examples. Theinvention is not to be interpreted as being limited to the Examples.

During the preparation of the Examples, generally:

-   -   a) Operations were carried out at ambient temperature, i.e. in        the range of about 17 to 30° C. and under an atmosphere of an        inert gas such as nitrogen unless otherwise stated;    -   b) Evaporations were carried out by rotary evaporation or        utilising Genevac equipment in vacuo and work-up procedures were        carried out after removal of residual solids by filtration;    -   c) Flash chromatography purifications were performed on an        automated Isco Combiflash Companion using Grace Resolve        prepacked silica columns, and (reverse phase flash) Isco        Combiflash Rf using RediSep Gold C18 columns;    -   d) Yields, where present, are not necessarily the maximum        attainable;    -   e) Structures of end-products of Formula (I) were confirmed by        nuclear magnetic resonance (NMR) spectroscopy, with NMR chemical        shift values measured on the delta scale. Proton magnetic        resonance spectra were determined using a Bruker Avance 700 (700        MHz), Bruker Avance 500 (500 MHz), Bruker 400 (400 MHz) or        Bruker 300 (300 MHz) instrument; ¹⁹F NMR were determined at 282        MHz or 376 MHz; ¹³C NMR were determined at 75 MHz or 100 MHz;        measurements were taken at around 20-30° C. unless otherwise        specified; the following abbreviations have been used: s,        singlet; d, doublet; t, triplet; q, quartet; m, multiplet; dd,        doublet of doublets; ddd, doublet of doublet of doublet; dt,        doublet of triplets; bs, broad signal;    -   f) End-products of Formula (I) were also characterised by mass        spectroscopy following liquid chromatography (LCMS), using a        HPLC system based on a Waters 2790/95 LC system with a 2996 PDA        and a 2000 amu ZQ single quadrupole mass spectrometer. The        solvents used were A=Water, B=Acetonitrile, C=50:50        acetonitrile:water 0.1% formic acid and D=50:50        acetonitrile:water 0.1% ammonium hydroxide. At a flow rate of        1.1 mL/min 5 L of sample was injected onto a 50×2.1 5 μm        Phenomenex Gemini NX column. The gradient ran from 95% A to 95%        B for 4.0 mins with a constant 5% infusion of C (for acid        analysis, D is used for base analysis). The flow was held at 95%        B for 0.5 mins before returning to start conditions. The Data        was acquired from 150 to 850 amu in both positive and negative        mode on the Mass Spectrometer and 220−320 nm on the PDA. LCMS        was also performed on a UPLC system utilising a Waters Acquity        Binary pump with sample manager, Acquity PDA and an SQD Mass        spectrometer. The solvents used were A1=0.1% formic acid (aq),        B1 0.1% formic acid in acetonitrile, A2=0.1% ammonium hydroxide        (aq) and B2 0.1% ammonium hydroxide in acetonitrile. At a flow        rate of 1 mL/min 1 L of sample was injected onto a 50×2.1 1.7 um        Waters BEH column (at 40° C.). The gradient ran from 97% A1 to        97% B1 over 1.30 mins before being held for 0.2 min and        returning to start conditions (substitute A1 and B1 for A2 and        B2 for base analysis). Data was acquired from 150-1000 amu in        positive and negative ion mode on the mass spectrometer and        245-320 amu on the PDA;    -   g) Intermediates were not generally fully characterised and        purity was assessed by thin layer chromatographic, mass        spectral, HPLC and/or NMR analysis;    -   h) The following abbreviations have been used: h=hour(s);        r.t.=room temperature (˜17-30° C.); conc.=concentrated;        FCC=flash column chromatography using silica;        AIBN=azobisisobutyronitrile; DCM=dichloromethane;        DIPEA=di-isopropyl ethylamine; DMA=N,N-dimethylacetamide;        DMF=N,N-dimethylformamide; DMSO=dimethylsulfoxide;        EDC=1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide; Et₂O=diethyl        ether; EtOAc=ethyl acetate; EtOH=ethanol;        HATU=1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium        3-oxid hexafluorophosphate; HOBT=hydroxybenzotriazole;        K₂CO₃=potassium carbonate; MeOH=methanol; MeCN=acetonitrile;        MgSO₄=anhydrous magnesium sulphate; Na₂SO₄=anhydrous sodium        sulphate; NBS=N-bromo succinimide; TFA=trifluoroacetic acid;        THE=tetrahydrofuran; sat.=saturated aqueous solution.

In a number of the examples below, a diastereomeric pair of compounds isdescribed. For example, the compounds of Example 1(a) and Example 1(b)represent a diastereomeric pair of compounds, formed as a mixture in theproduct of a single reaction and subsequently separated. In suchexamples, any assignment of stereochemistry is not absolute. By way ofillustration, Examples 1(a) and 1(b) relate to the (2S,3R) and (2R,3R)diastereomers of the named compound; however, it is not intended conveythat Example 1(a) is definitively assigned as the (2S,3R) diastereomerand Example 1(b) as the (2R,3R) diastereomer.

Example 1(a) and 1(b)(2S)-2-[3-(3-Fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-[3-(3-Fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

HATU (351 mg, 0.92 mmol) was added to[2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetyl]oxylithium(Intermediate 17, 170 mg, 0.71 mmol),N2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 187 mg, 0.71 mmol) and DIPEA (0.372 mL, 2.13 mmol) inN-methyl-2-pyrrolidinone (2 mL) and DMF (3 mL) at r.t. The resultingsolution was stirred at r.t. for 45 minutes. This solution was dilutedwith MeOH (15 mL) and passed through a 20 g SCX cartridge, flushing withMeOH followed by 1N NH₃ in MeOH to elute the product. The solvent wasevaporated under reduced pressure to yield crude product which wasdissolved in MeOH/DCM and evaporated onto silica gel. The residue waspurified by FCC (SiO₂, 0 to 12% MeOH in DCM). Pure fractions wereevaporated to dryness to afford the mixture of diastereoisomers as a gum(280 mg). The diastereoisomers were separated by preparative HPLC(Phenomenex Lux C4 column, 20 μm, 50 mm×250 mm, MeOH at 120 mL/min) togive:

First eluted isomer example 1(a),2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(65 mg, 23%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) 6 2.07 (1H, dq),2.23-2.32 (1H, m), 3.40-3.59 (3H, m), 3.71-3.78 (1H, m), 3.80-3.93 (2H,m), 4.14 (2H, dt), 4.37 (1H, dt), 4.87 (1H, s), 5.48 (1H, dtt), 6.43(1H, dd), 6.58 (1H, s), 6.80 (1H, d), 6.85 (1H, dd), 7.17 (1H, t), 7.32(1H, dd), 7.64 (1H, d), 8.47 (1H, dd), 12.10 (1H, s). m/z: ES⁺ [M+H]⁺485.

Second eluted isomer example 1(b)2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(68 mg, 24%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) 6 2.06 (1H, dd), 2.28(1H, td), 3.42-3.59 (3H, m), 3.75 (1H, dd), 3.8-3.93 (2H, m), 4.14 (2H,dt), 4.37 (1H, dt), 4.88 (1H, s), 5.48 (1H, ddd), 6.44 (1H, dd),6.57-6.61 (1H, m), 6.81 (1H, d), 6.86 (1H, dd), 7.18 (1H, t), 7.32 (1H,dd), 7.65 (1H, d), 8.47 (1H, dd), 12.12 (1H, s). m/z: ES⁺ [M+H]⁺ 485.

Example 2(a) and 2(b)(2S)-2-[3-(3,3-Difluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-[3-(3,3-Difluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

HATU (225 mg, 0.59 mmol) was added to2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-acetic acid(Intermediate 22, 117 mg, 0.46 mmol),N2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 120 mg, 0.46 mmol) and DIPEA (0.239 mL, 1.37 mmol) inN-methyl-2-pyrrolidinone (2 mL) and DMF (3 mL) at r.t. The resultingsolution was stirred at r.t. for 45 minutes. This solution was dilutedwith MeOH (15 mL) and passed through a 20 g SCX2 cartridge, flushingwith MeOH to remove impurities followed by a 1N solution of NH₃ in MeOHto elute the product. The solvent was evaporated under reduced pressureto yield crude product which was purified by FCC (SiO₂, 0 to 6% MeOH inDCM). Pure fractions were evaporated to afford the product as a mixtureof diastereoisomers as a gum (135 mg). The diastereoisomers wereseparated by preparative HPLC (Phenomenex Lux C2 column, 20 μm, 50mm×250 mm, MeOH/EtOH at 120 mL/min) to give:

First eluted isomer example 2(a)2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(55 mg, 41%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) 6 2.07 (1H, dt),2.22-2.32 (1H, m), 3.48 (1H, dd), 3.51-3.59 (2H, m), 3.74 (1H, dd), 4.25(4H, t), 4.37 (1H, dq), 4.89 (1H, s), 6.49-6.56 (1H, 1m), 6.67 (1H, d),6.81-6.91 (2H, m), 7.21 (1H, t), 7.31 (1H, dd), 7.64 (1H, d), 8.46 (1H,dd), 12.11 (1H, s). m/z: ES⁺ [M+H]⁺ 503.

Second eluted isomer example 2(b)2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(61 mg, 45%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) 2.00-2.11 (1H, m),2.22-2.31 (1H, m), 3.48 (1H, dd), 3.52-3.59 (2H, m), 3.74 (1H, dd), 4.25(4H, t), 4.36 (1H, dt), 4.88 (1H, s), 6.52 (1H, dd), 6.66 (1H, s),6.82-6.91 (2H, m), 7.21 (1H, t), 7.31 (1H, dd), 7.61 (1H, d), 8.47 (1H,dd), 12.11 (1H, s). m/z: ES⁺ [M+H]⁺ 503.

Example 3(a) and 3(b)(2S)-2-Methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-Methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

HATU (216 mg, 0.57 mmol) was added to2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetic acid (Intermediate25, 110 mg, 0.44 mmol),N2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 115 mg, 0.44 mmol) and DIPEA (0.229 mL, 1.31 mmol) inN-methyl-2-pyrrolidinone (2 mL) and DMF (3 mL) at r.t. The resultingsolution was stirred at r.t. for 45 minutes. This solution was dilutedwith MeOH (15 mL) and passed through a 20 g SCX2 cartridge, flushingwith MeOH to remove impurities followed by a 1N solution of NH₃ in MeOHto elute the product. The solvent was evaporated under reduced pressureto yield crude product. The crude product was dissolved in MeOH/DCM andevaporated down onto silica gel. The residue was purified by FCC (SiO₂,0 to 10% MeOH in DCM). Pure fractions were evaporated to dryness toafford the product as a mixture of diastereoisomers (140 mg) as a gum.The combined diastereoisomers were separated by preparative HPLC (Lux C4column, 20 μm, 50 mm×250 mm, 100% MeOH at 120 mL/min) to give:

First eluted isomer example 3(a)2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(74 mg, 15%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) 6 2.13 (1H, td),2.3-2.38 (1H, m), 3.30 (3H, s), 3.35 (3H, s), 3.54 (1H, dd), 3.58-3.67(4H, m), 3.81 (1H, dd), 4.05-4.12 (2H, m), 4.37 (1H, ddd), 4.44 (1H,dq), 4.92 (1H, s), 6.45 (1H, dd), 6.61 (1H, d), 6.82 (1H, d), 6.91 (1H,dd), 7.21 (1H, t), 7.38 (1H, dd), 7.71 (1H, d), 8.53 (1H, dd), 12.16(1H, s). m/z: ES⁺ [M+H]⁺ 497.

Second eluted isomer example 3(b)2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(67 mg, 14%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) 6 2.11 (1H, dq), 2.33(1H, dt), 3.29 (3H, s), 3.34 (3H, s), 3.54 (1H, dd), 3.58-3.66 (4H, m),3.80 (1H, dd), 4.05-4.12 (2H, m), 4.36 (1H, ddd), 4.4-4.47 (1H, m), 4.91(1H, s), 6.44 (1H, dd), 6.60 (1H, s), 6.82 (1H, d), 6.91 (1H, dd), 7.20(1H, t), 7.37 (1H, dd), 7.69 (1H, d), 8.53 (1H, dd), 12.15 (1H, s). m/z:ES⁺ [M+H]⁺ 497.

Example 4(a) and 4(b)(2S)-2-(3,5-Dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-(3,5-Dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

HATU (329 mg, 0.87 mmol) was added to2-(3,5-dimethoxyphenyl)-2-methoxy-acetic acid (Intermediate 28, 196 mg,0.87 mmol),N2-[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 6, 200 mg, 0.72 mmol) and DIPEA (0.252 mL, 1.44 mmol) inDMF (6 mL) at 21° C. under nitrogen. The resulting solution was stirredat 21° C. for 2 hours. The crude mixture was purified by ion exchangechromatography, using an SCX column. The desired product was eluted fromthe column using 1M NH₃ in MeOH and pure fractions were evaporated todryness to afford crude product which was purified by FCC (SiO₂, 0 to12% MeOH in DCM). Pure fractions were evaporated to dryness to afford abrown gum which was repurified by FCC (SiO₂, 0 to 8% MeOH in DCM). Purefractions were evaporated to dryness to afford the mixture ofdiastereoisomers as a yellow gum. The diastereoisomers were separated bypreparative HPLC (Lux C2 column, 20 μm, 50 mm×250 mm, 100% MeOH at 120mL/min) to give:

First eluted isomer example 4(a)2-(3,5-dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(21 mg, 6%). ¹H NMR (400 MHz, DMSO-d6, 27° C.) δ 1.98-2.10 (1H, m),2.18-2.31 (1H, m), 2.40 (3H, s), 3.30 (9H, s), 3.43 (1H, m), 3.47-3.61(2H, m), 3.72 (1H, m), 4.36 (1H, m), 4.87 (1H, s), 6.45 (1H, t), 6.62(2H, d), 6.81 (1H, d), 7.21 (1H, d), 7.64 (1H, s), 12.12 (1H, s). m/z:ES⁺ [M+H]⁺ 486.

Second eluted isomer example 4(b)2-(3,5-dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(18 mg, 5%). ¹H NMR (400 MHz, DMSO-d6, 27° C.) δ 1.98-2.10 (1H, m),2.18-2.31 (1H, m), 2.40 (3H, s), 3.30 (9H, s), 3.43 (1H, m), 3.47-3.61(2H, m), 3.72 (1H, m), 4.36 (1H, m), 4.87 (1H, s), 6.45 (1H, t), 6.62(2H, d), 6.81 (1H, d), 7.21 (1H, d), 7.64 (1H, s), 12.12 (1H, s). m/z:ES⁺ [M+H]⁺ 486.

Example 5(a) and 5(b)(2R)-3-Methoxy-2-(3-methoxyphenyl)-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]propanamideand(2S)-3-Methoxy-2-(3-methoxyphenyl)-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]propanamide

DIPEA (0.1 mL, 0.57 mmol), HATU (173.28 mg, 0.456 mmol) and3-methoxy-2-(3-methoxyphenyl)propanoic acid (Intermediate 29, 0.09 g,0.418 mmol) were added to a solution ofN2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 0.1 g, 0.38 mmol) in DMF (4 mL). The mixture wasstirred at r.t. for 18 h. This was diluted with water (5 mL), extractedinto DCM (10 mL), evaporated and purified by preparative HPLC (SunFireC18 column, 5 μm, 50 mm×19 mm, flow rate 25 mL/min). Decreasingly polarratios of water and MeCN containing 0.1% formic acid were used as amobile phase. Pure fractions were passed down an SCX cartridge washingwith methanol and then eluting with 2M NH₃ in MeOH. The basic fractionwas evaporated to dryness and the mixture of diastereoisomers wereseparated by SFC (Lux C1 column 5 μm, 21.2 mm×250 mm, at 40° C. at aflow rate of 50 mL/min, 50:50 MeOH:CO₂ containing 0.1% v/v NH₃) to give:

First eluted isomer example 5(a)3-methoxy-2-(3-methoxyphenyl)-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]propanamide(25.3 mg, 14.6%). ¹H NMR (400 MHz, DMSO-d6, 25° C.) δ 2.00-2.12 (1H, m),2.22-2.31 (1H, m), 3.25 (3H, s), 3.45-3.59 (4H, m), 3.71-3.79 (4H, m),3.97 (1H, t), 4.03-4.10 (1H, m), 4.31-4.43 (1H, m), 6.83-6.94 (4H, m),7.26 (1H, t), 7.33 (1H, dd), 7.68 (1H, d), 8.48 (1H, dd), 12.23 (1H, s).m/z: ES⁺ [M+H]⁺ 456.

Second eluted isomer example 5(b)3-methoxy-2-(3-methoxyphenyl)-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]propanamide(22.3 mg, 12.8%). ¹H NMR (400 MHz, DMSO-d6, 25° C.) δ 2.00-2.14 (1H, m),2.22-2.36 (1H, m), 3.25 (3H, s), 3.43-3.52 (2H, m), 3.52-3.61 (2H, m),3.69-3.78 (4H, m), 3.97 (1H, t), 4.01-4.10 (1H, m), 4.31-4.43 (1H, m),6.79-6.95 (4H, m), 7.26 (1H, t), 7.33 (1H, dd), 7.68 (1H, d), 8.47 (1H,dd), 12.23 (1H, s). m/z: ES⁺ [M+H]⁺ 456.

Example 6(a) and 6(b)(2S)-2-(3,5-Dimethoxyphenyl)-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-(3,5-Dimethoxyphenyl)-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

N2-[(3R)-1-Pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 0.2 g, 0.76 mmol) and2-(3,5-dimethoxyphenyl)-2-ethoxy-acetic acid (Intermediate 33, 0.18 g,0.76 mmol) were dissolved in DMF (2 mL) at r.t under N₂. The mixture wasstirred for 5 min before addition of DIPEA (0.34 mL, 1.943 mmol) andHATU (0.29 g, 0.76 mmol), then at r.t. for 1 h. The crude mixture waspassed through a 5 g SCX column washed with MeOH then eluted with 2N NH₃in MeOH. The basic fraction was purified by preparative HPLC (SunFireC18 column, 5 μm, 50 mm×19 mm, flow rate 25 mL/min). Decreasingly polarratios of water and MeCN containing 0.1% formic acid were used as amobile phase. Pure fractions were evaporated under reduced pressure andpassed through a 2 g SCX column washed with MeOH then eluted with 2N NH₃in MeOH. The basic fraction was evaporated under reduced pressure togive the product as a mixture of diastereoisomers as a beige foam. Thediastereoisomers were separated by SFC (Amy-C column, 5 μm, 20 mm×250mm, MeOH/CO₂ 45% containing NH₃ as modifier at 50 mL/min) to give:

First eluted isomer example 6(a)2-(3,5-dimethoxyphenyl)-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(70 mg, 19%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) δ 1.16 (3H, t), 2.04(1H, dq), 3.38-3.50 (3H, m), 3.51-3.58 (2H, m), 3.66-3.78 (7H, m), 4.35(1H, q), 4.96 (1H, s), 6.44 (1H, t), 6.62 (2H, d), 6.84 (1H, dd), 7.30(1H, dd), 7.67 (1H, d), 8.45 (1H, dd), 11.96 (1H, s). m/z: ES⁺ [M+H]⁺486.

Second eluted isomer example 6(b)2-(3,5-dimethoxyphenyl)-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(74 mg, 20%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) 1.18 (3H, t), 2.07 (1H,dt), 2.27 (1H, dt), 3.60-3.39 (5H, m), 3.79-3.70 (6H, m), 4.38 (1H, q),4.99 (1H, s), 6.47 (1H, t), 6.64 (2H, d), 6.88 (1H, dd), 7.34 (1H, dd),7.71 (1H, d), 8.48 (1H, dd), 12.15 (1H, s). m/z: ES⁺ [M+H]⁺ 486.

Example 7(a) and 7(b)(2S)-2-[4-Fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-N-[5-[(3R)-3-(pyridazin-3-ylamino)pyrrolidin-1-yl]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-[4-Fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-N-[5-[(3R)-3-(pyridazin-3-ylamino)pyrrolidin-1-yl]-1,3,4-thiadiazol-2-yl]acetamide

N2-[(3R)-1-Pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 108 mg, 0.41 mmol) and2-[4-fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-acetic acid(Intermediate 34, 110 mg, 0.41 mmol) were dissolved in DMF (3.5 mL) atr.t. under nitrogen. The mixture was stirred for 5 minutes beforeaddition of DIPEA (0.11 mL, 0.61 mmol) and HATU (186 mg, 0.49 mmol) thenstirred at r.t. overnight. The crude mixture was diluted with water (50mL) and then extracted into DCM (3×50 mL). The organic layer was dried(MgSO₄), filtered and then evaporated to give an orange gum which waspurified by preparative HPLC (SunFire C18 column, 5 μm, 50 mm×19 mm,flow rate 25 mL/min). Decreasingly polar ratios of water and MeCNcontaining 0.1% formic acid were used as a mobile phase. Fractionscontaining the desired product were combined, evaporated and passedthrough a 5 g SCX column washed with MeOH then eluted with 2 M NH₃ inMeOH. The basic fraction was evaporated to give the mixture ofdiastereoisomers. The diastereoisomers were then separated by SFC (LuxC3 column, 5 μm, 21.2 mm×250 mm, MeOH/CO₂ 30% containing NH₃ modifier,50 mL/min) to give:

First eluted isomer example 7(a)2-[4-fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-N-[5-[(3R)-3-(pyridazin-3-ylamino)pyrrolidin-1-yl]-1,3,4-thiadiazol-2-yl]acetamide(11 mg, 5%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) δ 1.94-2.12 (1H, m),2.19-2.36 (1H, m), 3.23 (3H, s), 3.27 (3H, s), 3.43-3.61 (3H, m),3.62-3.77 (3H, m), 4.07-4.19 (2H, m), 4.22-4.30 (1H, m), 4.32-4.43 (1H,m), 4.87 (1H, s), 6.66 (1H, dd), 6.73-6.84 (1H, m), 6.86 (1H, dd), 7.04(1H, dd), 7.32 (1H, dd), 7.70 (1H, d), 8.47 (1H, dd), 12.18 (1H, s).m/z: ES⁺ [M+H]⁺ 515.

Second eluted isomer example 7(b)2-[4-fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-N-[5-[(3R)-3-(pyridazin-3-ylamino)pyrrolidin-1-yl]-1,3,4-thiadiazol-2-yl]acetamide(11 mg, 5%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) δ 2.00-2.13 (1H, m),2.22-2.34 (1H, m), 3.23 (3H, s), 3.28 (3H, s), 3.42-3.60 (3H, m),3.63-3.80 (3H, m), 4.09-4.18 (2H, m), 4.24-4.33 (1H, m), 4.43-4.45 (1H,m), 4.87 (1H, s), 6.67 (1H, dd), 6.75-6.83 (1H, m), 6.87 (1H, dd), 7.05(1H, dd), 7.33 (1H, dd), 7.70 (1H, d), 8.48 (1H, dd), 12.18 (1H, s).m/z: ES⁺ [M+H]⁺ 515.

Example 8(a) and 8(b)(2S)-2-[4-Fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-[4-Fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

N2-[(3R)-1-Pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 100 mg, 0.38 mmol) and lithium2-[4-fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetate(Intermediate 38, 100 mg, 0.38 mmol) were dissolved in DMF (2.0 mL) atr.t under N₂. The mixture was stirred for 5 minutes before addition ofDIPEA (0.10 mL, 0.57 mmol) and HATU (173 mg, 0.46 mmol) then stirred atr.t. overnight. The crude mixture was passed through a 5 g SCX column,washed with MeOH, then eluted with 2M NH₃ in MeOH. The basic fractionwas evaporated to give an orange gum. The crude product was purified bypreparative HPLC (SunFire C18 column, 5 μm, 50 mm×19 mm at 25 mL/min).Decreasingly polar ratios of water and MeCN containing 0.1% formic acidwere used as a mobile phase. Fractions containing the product werecombined, evaporated and passed through a 5 g SCX column washed withMeOH then eluted with 2M NH₃ in MeOH. The basic fraction was evaporatedto give the mixture of diastereoisomers as an off-white solid (94 mg).The diastereoisomers were separated by preparative HPLC (Lux C4 column,20 μm, 50 mm×250 mm, MeOH/EtOH at 120 mL/min) to give:

First eluted isomer example 8(a)2-[4-fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(33 mg, 17%). ¹H NMR (400 MHz, DMSO, 31° C.) 2.03-2.13 (1H, m),2.24-2.33 (1H, m), 3.49 (1H, dd), 3.54-3.61 (2H, m), 3.75 (1H, dd), 3.97(2H, dd), 4.19-4.31 (2H, m), 4.35-4.43 (1H, m), 4.89 (1H, s), 5.36-5.56(1H, m), 6.71 (1H, dd), 6.8-6.89 (2H, m), 7.07 (1H, dd), 7.32 (1H, dd),7.65 (1H, d), 8.48 (1H, dd), 12.13 (1H, s). m/z: ES⁺ [M+H]⁺ 503.

Second eluted isomer example 8(b)2-[4-fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(34 mg, 18%). ¹H NMR (400 MHz, DMSO, 30° C.) 2.01-2.11 (1H, m),2.24-2.34 (1H, m), 3.47-3.61 (3H, m), 3.76 (1H, dd), 3.97 (2H, dd),4.18-4.32 (2H, m), 4.35-4.43 (1H, m), 4.88 (1H, s), 5.35-5.57 (1H, m),6.71 (1H, dd), 6.81-6.89 (2H, m), 7.07 (1H, dd), 7.33 (1H, dd), 7.64(1H, d), 8.48 (1H, d), 12.14 (1H, s), 21 assigned Hs: plus OMe protonsnot observed—obscured by solvent at 3.30 ppm. m/z: ES⁺ [M+H]⁺ 503.

Example 9(a) and 9(b)(2S)—N-[5-[[(3R)-1-(6-Fluoropyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetamideand(2R)—N-[5-[[(3R)-1-(6-Fluoropyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetamide

N2-[(3R)-1-(6-Fluoropyridazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 9, 0.11 g, 0.389 mmol) and[2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetyl]oxylithium(Intermediate 24, 0.12 g, 0.466 mmol) were dissolved in DMF (2 mL) atr.t under N₂. The mixture was stirred for 5 min before addition of DIPEA(0.34 mL, 1.943 mmol), and HATU (0.4 mL, 0.389 mmol), then at r.t. for 2h. The crude mixture was left to sit overnight, then passed through a 5g SCX column washed with MeOH then eluted with 2N NH₃ in MeOH. The basicfraction was evaporated to give an orange gum which was purified bypreparative HPLC (SunFire C18 column, 5 μm, 50 mm×19 mm, flow rate 25mL/min). Decreasingly polar ratios of water and MeCN containing 0.1%formic acid were used as a mobile phase. Pure fractions were combined,evaporated and passed through a 1 g SCX column washed with MeOH theneluted with 2N NH₃ in MeOH to give the mixture of diastereoisomers as anoff-white solid. The mixture of diastereoisomers was separated bypreparative chiral HPLC (Phenomenex Lux C4 column, 20 μm, 50 mm×250 mm,eluent MeOH at 120 mL/min) to give:

First eluted isomer example 9(a)N-[5-[[(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetamide(39.1 mg, 19.5%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) δ 2.07 (1H, dt),2.27 (1H, dt), 3.24 (3H, s), 3.28 (3H, s), 3.46 (1H, dd), 3.50-3.61 (4H,m), 3.73 (1H, dd), 4.00-4.07 (2H, m), 4.27-4.42 (2H, m), 4.84 (1H, s),6.39 (1H, dd), 6.55 (1H, s), 6.76 (1H, d), 7.12-7.20 (2H, m), 7.36 (1H,dd), 7.63 (1H, d), 12.16 (1H, s). m/z: ES⁺ [M+H]⁺ 515.

Second eluted isomer example 9(b)N-[5-[[(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetamide(42.8 mg, 21.4%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) δ 2.05 (1H, td),2.27 (1H, dt), 3.24 (3H, s), 3.28 (3H, s), 3.47 (1H, dd), 3.5-3.6 (4H,m), 3.73 (1H, dd), 3.99-4.07 (2H, m), 4.27-4.4 (2H, m), 4.83 (1H, s),6.39 (1H, dd), 6.55 (1H, d), 6.77 (1H, d), 7.12-7.2 (2H, m), 7.36 (1H,dd), 7.59 (1H, d), 12.17 (1H, s). m/z: ES⁺ [M+H]⁺ 515.

Example 10(a) and 10(b)(2S)-2-[3-(3-Fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-[3-(3-Fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

N2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 134 mg, 0.51 mmol) and lithium2-[3-(3-fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-acetate(Intermediate 40, 137 mg, 0.51 mmol) were dissolved in DMF (2 mL) at r.tunder N₂. The mixture was stirred for 5 minutes before addition of DIPEA(0.13 mL, 0.76 mmol) and HATU (232 mg, 0.61 mmol) then stirred at r.t.overnight. The crude mixture was passed through a 5 g SCX column, washedwith MeOH, then eluted with 2 M NH₃ in MeOH. The basic fraction wasevaporated to give an orange gum which was purified by preparative HPLC(SunFire C18 column, 5 μm, 50 mm×19 mm, flow rate 25 mL/min.Decreasingly polar ratios of water and MeCN containing 0.1% formic acidwere used as a mobile phase. Fractions containing the desired mass werecombined, evaporated and passed through a 5 g SCX column, washed withMeOH, then eluted with 2M NH₃ in MeOH. The basic fraction was evaporatedto give the mixture of diastereoisomers as an off-white solid (123 mg).The diastereoisomers were separated by preparative HPLC (Lux C4 column,20 μm, 50 mm×250 mm, 100% MeOH at 120 mL/min) to give:

First eluted isomer example 10(a)2-[3-(3-fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(45 mg, 17%). ¹H NMR (400 MHz, DMSO, 22° C.) 2.07 (1H, dq), 2.28 (1H,dt), 3.28 (3H, s), 3.49 (1H, dd), 3.52-3.59 (2H, m), 3.71 (3H, s), 3.74(1H, dd), 3.80-3.86 (1H, m), 3.86-3.92 (1H, m), 4.06-4.20 (2H, m), 4.37(1H, dt), 4.82 (1H, s), 5.47 (1H, dtd), 5.98 (1H, t), 6.13-6.21 (1H, m),6.39-6.45 (1H, m), 6.86 (1H, dd), 7.33 (1H, dd), 7.67 (1H, d), 8.48 (1H,dd), 12.13 (1H, s). m/z: ES⁺ [M+H]⁺ 515.

Second eluted isomer example 10(b)2-[3-(3-fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(46 mg, 17%). ¹H NMR (400 MHz, DMSO, 22° C.) 2.06 (1H, dq), 2.27 (1H,dt), 3.28 (3H, s), 3.49 (1H, dd), 3.53-3.59 (2H, m), 3.71 (3H, s), 3.75(1H, dd), 3.80-3.86 (1H, m), 3.87-3.92 (1H, m), 4.13 (2H, dt), 4.37 (1H,dt), 4.81 (1H, s), 5.47 (1H, dtt), 5.97 (1H, t), 6.13-6.23 (1H, m),6.38-6.45 (1H, m), 6.87 (1H, dd), 7.33 (1H, dd), 7.64 (1H, d), 8.48 (1H,dd), 12.15 (1H, s). m/z: ES⁺ [M+H]⁺ 515.

Example 11(a) and 11(b)(2S)-2-Methoxy-2-[3-methoxy-5-(trifluoromethoxy)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-methoxy-2-[3-methoxy-5-(trifluoromethoxy)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

N2-[(3R)-1-Pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 100 mg, 0.38 mmol) and2-methoxy-2-[3-methoxy-5-(trifluoromethoxy)phenyl]acetic acid(Intermediate 44, 191 mg, 0.68 mmol) were dissolved in DMF (3.5 mL) atr.t under N₂. The mixture was stirred for 5 minutes before addition ofDIPEA (0.1 mL, 0.57 mmol) and HATU (173 mg, 0.46 mmol) then stirred atr.t. overnight. The crude mixture was diluted with water (50 mL) andextracted into DCM (50 mL×2). The organic layer was dried (MgSO₄),filtered and evaporated to afford an orange gum which was purified bypreparative HPLC (SunFire C18 column, 5 μm, 50 mm×19 mm, flow rate 25mL/min). Decreasingly polar ratios of water and MeCN containing 0.1%formic acid were used as a mobile phase. Fractions containing thedesired mass were combined, evaporated and passed through a 5 g SCXcolumn washed with MeOH then eluted with 2M NH₃ in MeOH. The basicfraction was evaporated to give a yellow solid. The mixture ofdiastereoisomers was separated by preparative HPLC (Phenomenex Lux C2column, 20 μm, 50 mm×250 mm, using a 50/50 mixture of EtOH/MeOH aseluents at 120 mL/min) to give:

First eluted isomer example 11(a)2-methoxy-2-[3-methoxy-5-(trifluoromethoxy)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(9.1 mg, 4.5%). ¹H NMR (400 MHz, CDCl₃, 30° C.) δ 2.28 (1H, dd),2.35-2.47 (1H, m), 3.48 (3H, s), 3.56-3.71 (2H, m), 3.73-3.83 (4H, m),3.87 (1H, dd), 4.52 (1H, s), 4.84 (1H, s), 6.25 (1H, s), 6.56 (1H, dd),6.71-6.76 (1H, m), 6.89-6.95 (2H, m), 7.11 (1H, dd), 8.48 (1H, dd). m/z:ES⁺ [M+H]⁺ 526.

Second eluted isomer example 11(b)2-methoxy-2-[3-methoxy-5-(trifluoromethoxy)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(10.2 mg 5.1%). ¹H NMR (400 MHz, CDCl₃, 30° C.) δ 2.20-2.31 (1H, m),2.35-2.47 (1H, m), 3.47 (3H, s), 3.56-3.78 (3H, m), 3.80 (3H, s),3.84-3.93 (1H, m), 4.50 (1H, s), 4.84 (1H, s), 6.14 (1H, s), 6.57 (1H,d), 6.73 (1H, s), 6.92 (2H, d), 7.12 (1H, dd), 8.49 (1H, s). m/z: ES⁺[M+H]⁺ 526.

Example 12(a) and 12(b)(2S)-2-Methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-Methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

N2-[(3R)-1-(6-Methylpyridazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 6, 0.11 g, 0.389 mmol) and[2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetyl]oxylithium(Intermediate 24, 0.12 g, 0.466 mmol) were dissolved in DMF (2 mL) atr.t under N₂. The mixture was stirred for 5 min before addition of DIPEA(0.34 mL, 1.943 mmol), and HATU (0.4 mL, 0.389 mmol), then at r.t. for2h. The crude mixture was passed through a 5 g SCX column washed withMeOH, then eluted with 2N NH₃ in MeOH. The basic fraction was evaporatedand purified by preparative HPLC (SunFire C18 column, 5 μm, 50 mm×19 mmat 25 mL/min). Decreasingly polar ratios of water and MeCN containing0.1% formic acid were used as a mobile phase. Pure fractions werecombined, evaporated and passed through a 1 g SCX column washed withMeOH then eluted with 2N NH₃ in MeOH to give the mixture ofdiastereoisomers as an off-white solid (98 mg). The diastereoisomerswere separated by preparative HPLC (Lux C4 column, 20 μm, 50 mm×250 mm,100% MeOH at 120 mL/min) to give:

First eluted isomer example 12(a)2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(34 mg, 18%). ¹H NMR (400 MHz, DMSO, 30° C.) 2.00-2.11 (1H, m), 2.30(1H, dt), 2.41 (3H, s), 3.25 (3H, s), 3.45 (1H, dd), 3.49-3.62 (4H, m),3.73 (1H, dd), 4.04 (2H, t), 4.28-4.41 (2H, m), 4.86 (1H, s), 6.40 (1H,d), 6.55 (1H, s), 6.77 (1H, d), 6.82 (1H, d), 7.15 (1H, t), 7.22 (1H,d), 7.62 (1H, d), 12.06 (1H, s); m/z: ES⁺ [M+H]⁺ 511.

Second eluted isomer example 12(b)2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(38 mg, 19%). ¹H NMR (400 MHz, DMSO, 30° C.) 2.05 (1H, dt), 2.27 (1H,dt), 2.42 (3H, s), 3.25 (3H, s), 3.46 (1H, dd), 3.5-3.61 (4H, m), 3.74(1H, dd), 4.04 (2H, t), 4.28-4.41 (2H, m), 4.85 (1H, s), 6.39 (1H, dd),6.55 (1H, s), 6.77 (1H, d), 6.82 (1H, d), 7.15 (1H, t), 7.22 (1H, d),7.58 (1H, d), 12.09 (1H, s). m/z: ES⁺ [M+H]⁺ 511.

Example 13(a) and 13(b)(2S)-2-Methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-Methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

2-Methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetyl]oxylithium(Intermediate 24, 0.12 g, 0.475 mmol) andN2-[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 12, 0.13 g, 0.475 mmol) were weighed into a round bottomedflask. DMF (3 mL) and DIPEA (0.15 g, 1.187 mmol) were added followed byHATU (0.18 g, 0.475 mmol) and the resultant solution was allowed to stirat r.t. under N₂ for 15 h. The reaction mixture was added to an SCXcartridge and washed with MeOH then eluted with 2M NH₃ in MeOH. Thebasic fractions were evaporated under reduced pressure and the residuepurified by preparative HPLC (SunFire C18 column, 5 μm, 50 mm×19 mm at25 mL/min). Decreasingly polar ratios of water and MeCN containing 0.1%formic acid were used as a mobile phase. Fractions containing thedesired mass were evaporated under reduced pressure to give the mixtureof diastereoisomers as an orange gum. The diastereoisomers was separatedby preparative chiral HPLC (Lux C1 column, 5 μm, 21 mm×250 mm, eluent20:80 heptane:EtOH at 21 mL/min containing 0.1% NH₃) to give:

First eluted isomer example 13(a)2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(55.2 mg, 16.5%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ 2.09 (1H, s), 2.30(1H, d), 3.24 (3H, s), 3.29 (3H, s), 3.53-3.90 (6H, m), 4.03 (2H, tt),4.27-4.42 (2H, m), 4.86 (1H, s), 6.40 (1H, ddd), 6.55 (1H, s), 6.74-6.80(1H, m), 7.15 (1H, t), 7.70 (1H, d), 8.32 (1H, d), 8.61 (1H, d), 12.17(1H, s). m/z: ES⁺ [M+H]⁺ 498.

Second eluted isomer example 13(b)2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(54.2 mg, 13.7%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ 1.98-2.08 (1H, m),2.19-2.28 (1H, m), 3.18 (3H, s), 3.22 (3H, s), 3.47-3.84 (6H, m),3.93-4.02 (2H, m), 4.21-4.36 (2H, m), 4.80 (1H, s), 6.34 (1H, ddd), 6.49(1H, s), 6.67-6.74 (1H, m), 7.09 (1H, t), 7.64 (1H, d), 8.26 (1H, d),8.55 (1H, d), 12.11 (1H, s). m/z: ES⁺ [M+H]⁺ 498.

Example 14(a) and 14(b)(2S)-2-[3-(3,3-Difluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-[3-(3,3-Difluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

Into a round bottomed flask was weighedN2-[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 12, 0.06 g, 0.232 mmol),[2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-acetyl]oxylithium(Intermediate 21, 0.06 g, 0.232 mmol). DMF (2 mL) and DIPEA (0.1 mL,0.579 mmol) were added followed by HATU (0.09 g, 0.232 mmol) and theresultant solution was allowed to stir at r.t. for 15 hours. Thereaction mixture was added to an SCX cartridge and washed with MeOH theneluted with 2M NH₃ in MeOH. The basic fractions were evaporated underreduced pressure and the residue was purified by preparative HPLC(SunFire C18 column, 5 μm, 50 mm×19 mm at 25 mL/min). Decreasingly polarratios of water and MeCN containing 0.1% formic acid were used as amobile phase. The solvent was removed under reduced pressure to give themixture of diastereoisomers as a cream solid. The diastereoisomers wereseparated by chiral preparative HPLC (Amy C column, 5 μm, 20 mm×250 mm,eluent 20:80 heptane:EtOH containing 0.1% NH₃ at 21 mL/min) to give:

First eluted isomer example 14(a)2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(22.1 mg, 18.9%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ 2.02-2.12 (1H, m),2.23-2.34 (1H, m), 3.29 (3H, s), 3.77 (4H, s), 4.20-4.42 (5H, m), 4.89(1H, s), 6.49-6.57 (1H, m), 6.67 (1H, t), 6.86-6.92 (1H, m), 7.22 (1H,t), 7.67 (1H, d), 8.32 (1H, d), 8.61 (1H, d), 12.19 (1H, s). m/z:ES+[M+H]⁺ 504.

Second eluted isomer example 14(b)2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(20.8 mg, 17.8%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ 2.02-2.13 (1H, m),2.24-2.35 (1H, m), 3.29 (3H, s), 3.77 (4H, s), 4.20-4.42 (5H, m), 4.89(1H, s), 6.50-6.56 (1H, m), 6.65-6.69 (1H, m), 6.86-6.91 (1H, m), 7.22(1H, t), 7.69 (1H, d), 8.31 (1H, d), 8.61 (1H, d), 12.19 (1H, s). m/z:ES⁺ [M+H]⁺ 504.

Example 15(a) and 15(b)(2S)-2-[3-(3-Fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-[3-(3-Fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

Into a round bottomed flask was weighedN2-[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 12, 0.05 g, 0.17 mmol),[2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetyl]oxylithium(Intermediate 17, 0.04 g, 0.17 mmol). DMF (2 mL) and HATU (0.06 g, 0.17mmol) were added followed by DIPEA (0.07 mL, 0.426 mmol) and theresultant solution was allowed to stir at r.t. for 15 h. The reactionmixture was added to an SCX cartridge and washed with MeOH then elutedwith 2M NH₃ in MeOH. The basic fractions were evaporated under reducedpressure and the residue was purified by preparative HPLC (SunFire C18column, 5 μm, 50 mm×19 mm at 25 mL/min). Decreasingly polar ratios ofwater and MeCN containing 0.1% formic acid were used as a mobile phase.The solvent was removed under reduced pressure to give the mixture ofdiastereoisomers as a cream solid. The diastereoisomers was separated bypreparative chiral HPLC (Amy-C column, 5 μm, 20 mm×250 mm, eluent 20:80heptane:EtOH at 21 mL/min containing 0.1% NH₃) to give:

First eluted isomer example 15(a)2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(17.9 mg, 21.7%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ 2.04-2.13 (1H, m),2.23-2.35 (1H, m), 3.28 (3H, s), 3.47-3.94 (6H, m), 4.06-4.20 (2H, m),4.35 (1H, s), 4.84 (1H, s), 5.55 (1H, tt), 6.39-6.46 (1H, m), 6.58 (1H,t), 6.80 (1H, dt), 7.16 (1H, t), 7.62 (1H, s), 8.31 (1H, d), 8.60 (1H,d), 12.18 (1H, s). m/z: ES⁺ [M+H]⁺ 486.

Second eluted isomer example 15(b)2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(19 mg, 23%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ 2.03-2.13 (1H, m),2.23-2.34 (1H, m), 3.28 (s, 3H), 3.50-3.93 (6H, m), 4.08-4.21 (2H, m),4.36 (1H, s), 4.87 (1H, s), 5.38-5.58 (1H, m), 6.41-6.46 (1H, m), 6.58(1H, t), 6.77-6.83 (1H, m), 7.17 (1H, t), 7.68 (1H, d), 8.32 (1H, d),8.61 (1H, d), 12.18 (1H, s). m/z: ES⁺ [M+H]⁺ 486.

Example 16(a) and 16(b)(2S)-2-(3,5-Dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-(3,5-Dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

DIPEA (0.1 mL, 0.568 mmol) was added to2-(3,5-dimethoxyphenyl)-2-methoxy-acetic acid (Intermediate 28, 0.13 g,0.568 mmol) andN2-[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 12, 0.15 g, 0.568 mmol) in DMF (3 mL). HATU (0.22 g, 0.568mmol) was added to the solution and the reaction mixture was allowed tostir at room temperature under nitrogen for 24 h. The solvent wasremoved under reduced pressure to provide a dark orange oil which wasdissolved in DCM, absorbed onto silica and purified by FCC (SiO₂, 1-10%MeOH in DCM). The pure fractions were combined and solvent was removedunder reduced pressure to provide a dark orange oil which was furtherpurified by preparative HPLC (SunFire C18 column, 5 μm, 50 mm×19 mm,flow rate 25 mL/min). Decreasingly polar ratios of water and MeCNcontaining 0.1% formic acid were used as a mobile phase. Pure fractionswere combined, evaporated under reduced pressure and passed through a 2g SCX column washed with MeOH then eluted with 2N NH₃ in MeOH. The basicfraction was evaporated to dryness to afford the mixture ofdiastereoisomers as an off-white foam (132 mg). The diastereoisomerswere separated by preparative HPLC (Lux C4 column, 5 μm, 20 mm×250 mm,eluent MeOH containing diethanolamine modifier at 21 mL/min). Theseparate diastereoisomers were dissolved in DCM, washed with water andthe organic layer evaporated to give:

First eluted isomer example 16(a)2-(3,5-dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(23.2 mg, 8.6%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) δ 2.09 (1H, d) 2.30(1H, dd), 3.30 (3H, d), 3.74 (6H, s), 3.55-3.91 (4H, m), 4.38 (1H, s),4.89 (1H, s), 6.47 (1H, t), 6.64 (2H, d), 7.71 (1H, d), 8.32 (1H, d),8.61 (1H, d), 12.17 (1H, s). m/z: ES⁺ [M+H]⁺ 472.

Second eluted isomer example 16(b)2-(3,5-dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(28.3 mg, 10.5%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) 2.12 (1H, s), 2.31(1H, d), 3.33 (3H, s), 3.60-3.73 (3H, m), 3.77 (6H, s), 3.80-3.89 (1H,m), 4.40 (1H, s), 4.92 (1H, s), 6.50 (1H, t), 6.67 (2H, d), 7.73 (1H,d), 8.35 (1H, d), 8.64 (1H, d), 12.19 (1H, s). m/z: ES⁺ [M+H]⁺ 472.

Example 17(a) and 17(b)(2S)-[3-(Difluoromethoxy)phenyl]-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-[3-(Difluoromethoxy)phenyl]-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

2-[3-(Difluoromethoxy)phenyl]-2-ethoxy-acetic acid (Intermediate 47,0.14 g, 0.57 mmol) andN2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 0.15 g, 0.57 mmol) were weighed into a round bottomedflask. DMF (3 mL) and DIPEA (0.18 g, 1.424 mmol) were added followed byHATU (0.22 g, 0.57 mmol) and the resultant solution was allowed to stirat r.t. under N₂ for 3 h. The solvent was removed under reducedpressure. The residual gum was dissolved in DCM, absorbed onto silicaand purified by FCC (SiO₂, 1 to 8% MeOH in DCM). Evaporation of the purefractions under reduced pressure gave a yellow gum that was separated bypreparative chiral SFC (Amy-C column, 5 μm, 20 mm×250 mm, eluentMeOH/CO₂ containing 40% NH₃ as a modifier. The flow rate was 50 mL/minat a wavelength of 210 nm) to give:

First eluted isomer example 17(a)[3-(difluoromethoxy)phenyl]-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(23 mg, 8.2%). ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ 1.19 (3H, t),2.02-2.10 (1H, m), 2.21-2.36 (1H, m), 3.37-3.63 (5H, m), 3.74 (1H, dd),4.40-4.36 (1H, m), 5.12 (1H, s), 6.87 (1H, dd), 7.16 (1H, dd), 7.25 (1H,t), 7.28-7.29 (1H, m), 7.30-7.38 (2H, m), 7.45 (1H, t), 7.72 (1H, d),8.48 (1H, dd), 12.27 (1H, s). m/z: ES⁺ [M+H]⁺ 492.

Second eluted isomer example 17(b)[3-(difluoromethoxy)phenyl]-2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(0.025 g, 8.9%).

¹H NMR (400 MHz, DMSO-d₆, 30° C.) δ ¹H NMR (400 MHz, DMSO-d6) δ 1.19(3H, t), 2.03-2.10 (1H, m), 2.24-2.32 (1H, m), 3.38-3.62 (5H, m), 3.74(1H, dd), 4.36-4.40 (1H, m), 5.12 (1H, s), 6.86 (1H, dd), 7.16 (1H, t),7.24 (1H, t), 7.26-7.38 (3H, m), 7.45 (1H, t), 7.72 (1H, d), 8.48 (1H,dd), 12.27 (1H, s). m/z: ES⁺ [M+H]⁺ 492.

Example 18(a) and 18(b)(2S)-2-Methoxy-2-[3-(oxetan-3-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-Methoxy-2-[3-(oxetan-3-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

HATU (520 mg, 1.37 mmol) was added toN2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 300 mg, 1.14 mmol),2-methoxy-2-[3-(oxetan-3-yl)phenyl]acetic acid (Intermediate 48, 304 mg,1.37 mmol) and DIPEA (0.198 mL, 1.14 mmol) in DMF (8 mL) at 21° C. underN₂. The resulting solution was stirred at 21° C. for 2 hours. The crudeproduct was purified by ion exchange chromatography, using an SCXcolumn. The desired product was eluted from the column using 1M NH₃ inMeOH and pure fractions were evaporated. The crude product was purifiedby FCC (SiO₂, 0 to 15% 1M NH₃ in MeOH in EtOAc). Pure fractions wereevaporated to dryness to afford a solid. The crude product was purifiedby preparative HPLC (Waters XBridge OBD C18 column, 5 μm, 30 mm×100 mm).Decreasingly polar mixtures of water containing 1% NH₄₀H and MeCN wereused as the mobile phase. Fractions containing the desired compound wereevaporated to dryness to yield the product as a mixture ofdiastereoisomers. The diastereoisomers were separated by preparativeHPLC (Phenomenex Lux C4 column, 20 μm, 50 mm×250 mm, MeOH/IPA 50/50 at120 mL/min) to give:

First eluted isomer example 18(a)2-methoxy-2-[3-(oxetan-3-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(45 mg, 8%). ¹H NMR (400 MHz, DMSO, 30° C.) δ 2.12 (1H, td), 2.23-2.40(1H, m), 3.38 (3H, s), 3.46-3.69 (3H, m), 3.74-3.86 (1H, m), 4.26-4.35(1H, m), 4.41-4.50 (1H, m), 4.65 (2H, ddd), 4.90-5.11 (3H, m), 6.91 (1H,dd), 7.27-7.49 (4H, m), 7.57 (1H, s), 7.70 (1H, d), 8.53 (1H, dd), 12.24(1H, s). m/z: ES⁺ [M+H]⁺ 468.

Second eluted isomer example 18(b)2-methoxy-2-[3-(oxetan-3-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(38 mg, 7%). ¹H NMR (400 MHz, DMSO, 30° C.) δ 2.12 (1H, td), 2.23-2.40(1H, m), 3.38 (3H, s), 3.46-3.69 (3H, m), 3.74-3.86 (1H, m), 4.26-4.35(1H, m), 4.41-4.50 (1H, m), 4.65 (2H, ddd), 4.90-5.11 (3H, m), 6.91 (1H,dd), 7.27-7.49 (4H, m), 7.57 (1H, s), 7.70 (1H, d), 8.53 (1H, dd), 12.24(1H, s). m/z: ES⁺ [M+H]⁺ 468.

Example 19(a) and 19(b)(2S)-2-Ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-[3-(trifluoromethoxy)phenyl]acetamideand(2R)-2-Ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-[3-(trifluoromethoxy)phenyl]acetamide

DIPEA (0.15 mL, 0.85 mmol), HATU (260 mg, 0.68 mmol) and2-ethoxy-2-[3-(trifluoromethoxy)phenyl]acetic acid (Intermediate 51, 180mg, 0.68 mmol) were added to a solution ofN2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 150 mg, 0.57 mmol) in DMF (4 mL). The mixture wasstirred at r.t. for 18 h. This was then diluted with water (5 mL) andthen extracted into DCM (10 mL), evaporated and purified by preparativeHPLC (XBridge OBD C18 column, 5 μm, 50 mm×19 mm, flow rate was 25mL/min). Decreasingly polar ratios of water and MeCN containing 0.3 mL/LNH₄OH were used as a mobile phase. Pure fractions were evaporated andpassed through an SCX cartridge washing with MeOH and then eluting with2M NH₃ in MeOH. The basic fraction was evaporated and dried in vacuo togive the product as a mixture of diastereoisomers. The diastereoisomerswere then separated by HPLC (Lux C4 column, 5 μm, 20 mm×250 mm, MeOHcontaining NH₃ modifier, 21 mL/min) to give:

First eluted isomer example 19(a)2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-[3-(trifluoromethoxy)phenyl]acetamide,(46 mg, 16%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) δ 1.19 (3H, t),1.97-2.15 (1H, m), 2.18-2.36 (1H, m), 3.40-3.65 (5H, m), 3.65-3.81 (1H,m), 4.32-4.46 (1H, m), 5.17 (1H, s), 6.87 (1H, dd), 7.28-7.40 (2H, m),7.40-7.59 (3H, m), 7.74 (1H, d), 8.48 (1H, dd), 12.32 (1H, s). m/z: ES⁺[M+H]⁺ 510.

Second eluted isomer example 19(b)2-ethoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-[3-(trifluoromethoxy)phenyl]acetamide(39 mg, 14%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) δ 1.19 (3H, t), 2.06(1H, m), 2.20-2.35 (1H, m), 3.39-3.63 (5H, m), 3.74 (1H, m), 4.34-4.44(1H, m), 5.17 (1H, s), 6.88 (1H, dd), 7.28-7.41 (2H, m), 7.42-7.58 (3H,m), 7.73 (1H, d), 8.48 (1H, d), 12.32 (1H, s). m/z: ES⁺ [M+H]⁺ 510.

Example 20(a) and 20(b)(2R)-2-[4-Fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2S)-2-[4-Fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

DIPEA (0.12 mL, 0.71 mmol), HATU (216 mg, 0.568 mmol) and2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-acetic acid(Intermediate 52, 0.13 g, 0.474 mmol) were added to a solution ofN2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 0.12 g, 0.474 mmol) in DMF (4 mL). The mixture wasstirred at r.t. for 18 h. This was then diluted with water (5 mL),extracted into DCM (10 mL), evaporated and purified by preparative HPLC(SunFire C18 column, 5 μm, 50 mm×19 mm at 25 mL/min). Decreasingly polarratios of water and MeCN containing 0.1% formic acid were used as amobile phase. Pure fractions were passed down an SCX cartridge washingwith MeOH and then eluting with 2M NH₃ in MeOH. The basic fraction wasevaporated. An impurity was still seen so the material was re-purifiedby preparative HPLC (XBridge OBD C18 column, 5 μm, 50 mm×19 mm, flowrate 25 mL/min). Decreasingly polar ratios of water and MeCN containing0.3 mL/L NH₄OH were used as a mobile phase. Pure fractions wereevaporated to dryness and the mixture of diastereoisomers was separatedby preparative chiral HPLC (Phenomenex Lux C2 column, 20 μm, 50 mm×250mm, eluent MeOH at 110 mL/min) to give:

First eluted isomer example 20(a)2-[4-Fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(18.4 mg, 7.5%). ¹H NMR (400 MHz, DMSO-d6, 22° C.) 2.06 (1H, dq), 2.28(1H, dt), 3.34 (3H, s), 3.45-3.60 (3H, m), 3.75 (1H, dd), 4.34-4.42 (1H,m), 5.03 (1H, s), 6.87 (1H, dd), 7.33 (1H, dd), 7.51-7.57 (2H, m),7.60-7.69 (2H, m), 8.48 (1H, dd), 12.35 (1H, s). m/z: ES⁺ [M+H]⁺ 514.

Second eluted isomer example 20(b)2-[4-Fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide(19.2 mg, 7.9%). ¹H NMR (400 MHz, DMSO-d6, 22° C.) 2.07 (1H, dd), 2.28(1H, dt), 3.49 (1H, d), 3.53-3.60 (2H, m), 3.74 (1H, dd), 4.33-4.43 (1H,m), 5.04 (1H, s), 6.87 (1H, d), 7.33 (1H, dd), 7.52-7.57 (2H, m), 7.63(1H, d), 7.68 (1H, d), 8.48 (1H, d), 12.35 (1H, s). m/z: ES⁺ [M+H]⁺ 514.

Example 21(a) and 21(b)(2S)-2-(3,5-Dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-(3,5-Dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

HATU (433 mg, 1.14 mmol) was added to2-(3,5-dimethoxyphenyl)-2-methoxy-acetic acid (Intermediate 28, 215 mg,0.95 mmol),N2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 250 mg, 0.95 mmol) and DIPEA (0.332 mL, 1.90 mmol) inDMF (8 mL) at 21° C. under nitrogen. The resulting solution was stirredat 21° C. for 2 hours. The crude product was purified by ion exchangechromatography, using an SCX column. The desired product was eluted fromthe column using 1M NH₃/MeOH and pure fractions were evaporated todryness to afford crude product. The crude product was purified by flashsilica chromatography, (elution gradient 0 to 12% 1M NH₃/MeOH in DCM).Pure fractions were evaporated to dryness to afford a beige foam. LCMSshowed a 9% chemical impurity. The crude product was further purified byflash silica chromatography, (elution gradient 0 to 8% MeOH in DCM).Pure fractions were evaporated to dryness to afford the product as amixture of diastereoisomers. (180 mg)

The diastereoisomers were separated by preparative HPLC (Luc C4 column,20 μm silica, 50 mm diameter, 250 mm length, 100% MeOH at 120 ml/min).The separate diastereoisomers were dissolved in DCM, washed with waterand the organic layer evaporated to give:

Example 21(a) as the first eluted isomer (solid, 74 mg, 16%). ¹H NMR(400 MHz, DMSO, 27° C.) 62.06 (1H, m), 2.21-2.38 (1H, m), 3.30 (9H, d),3.41-3.62 (3H, m), 4.28-4.47 (1H, m), 4.87 (1H, s), 6.46 (1H, t), 6.63(2H, s), 6.85 (1H, d), 7.31 (1H, dd), 7.65 (1H, d), 8.47 (1H, dd), 12.13(1H, s). m/z: ES⁺ [M+H]⁺ 472.

Example 21(b) as the second eluted isomer (solid, 75 mg, 17%). ¹H NMR(400 MHz, DMSO, 27° C.) 62.06 (1H, m), 2.21-2.38 (1H, m), 3.30 (9H, d),3.41-3.62 (3H, m), 4.28-4.47 (1H, m), 4.87 (1H, s), 6.46 (1H, t), 6.63(2H, s), 6.85 (1H, d), 7.31 (1H, dd), 7.65 (1H, d), 8.47 (1H, dd), 12.13(1H, s). m/z: ES⁺ [M+H]⁺ 472.

Example 22(a) and 22(b)(2S)-2-ethoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamideand(2R)-2-ethoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide

HATU (0.13 g, 0.332 mmol) andN2-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 1, 0.07 g, 0.276 mmol) were dissolved in DMF (2 mL) at r.tunder N₂. The mixture was stirred for 5 min before addition of DIPEA(0.07 mL, 0.414 mmol) and lithium2-ethoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetate (Intermediate 56,0.07 g, 0.276 mmol) then allowed to stir at r.t. overnight. The crudemixture was passed through a 5 g SCX column washed with MeOH then elutedwith 2N NH₃ in MeOH. The basic fraction was evaporated to give theimpure product as an orange gum which was purified by preparative HPLC(SunFire C18 column, 5 μm, 50 mm×19 mm at 25 mL/min). Decreasingly polarratios of water and MeCN containing 0.1% formic acid were used as amobile phase. Fractions containing the desired mass were combined,evaporated and passed through a 5 g SCX column, washing with MeOH, theneluting with 2N NH₃ in MeOH. The solvent was removed in vacuo to give amixture of diastereoisomers which were separated by preparative chiralHPLC. (Phenomenex Lux C4 column, 20 μm silica, 50 mm diameter, 250 mmlength), EtOH/MeOH 50/50 at 120 mL/min. Fractions containing the desiredcompounds were evaporated to dryness to give:

Example 22(a) as the first eluted isomer (solid, 21.4 mg, 15%).

Example 22(b) as the second eluted isomer (solid, 20.4 mg, 14%).

ADDITIONAL EXAMPLES

The compounds of the following Examples were prepared in a similarfashion to the Examples above.

Example no. Name MS data 232-(3,5-dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)- m/z: ES+1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4- [M + H]+thiadiazol-2-yl]acetamide 472 242-methoxy-2-[3-(oxetan-3-yl)phenyl]-N-[5-[[(3R)- m/z (ES+),1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4- [M + H]+ =thiadiazol-2-yl]acetamide 469 252-methoxy-2-[3-(oxetan-3-yl)phenyl]-N-[5-[[(3R)- m/z (ES+),1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4- [M + H]+ =thiadiazol-2-yl]acetamide 469 262-(3,5-dimethoxyphenyl)-N-[5-[[(3R)-1-(6- m/z (ES+),fluoropyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4- [M + H]+ =thiadiazol-2-yl]-2-methoxy-acetamide 490 272-[3-(3-fluoroazetidin-1-yl)phenyl]-N-[5-[[(3R)-1- m/z: ES+(6-fluoropyridazin-3-yl)pyrrolidin-3-yl]amino]- [M + H]+1,3,4-thiadiazol-2-yl]-2-methoxy-acetamide 503 282-[3-(3-fluoroazetidin-1-yl)phenyl]-N-[5-[[(3R)-1- m/z: ES+(6-fluoropyridazin-3-yl)pyrrolidin-3-yl]amino]- [M + H]+1,3,4-thiadiazol-2-yl]-2-methoxy-acetamide 503 292-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-ethoxy- m/z: ES+N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3- [M + H]+yl]amino]-1,3,4-thiadiazol-2-yl]acetamide 517 302-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-ethoxy- m/z: ES+N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3- [M + H]+yl]amino]-1,3,4-thiadiazol-2-yl]acetamide 517 312-ethoxy-2-[3-(3-fluoroazetidin-1-yl)phenyl]-N-[5- m/z: ES+[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]- [M + H]+1,3,4-thiadiazol-2-yl]acetamide 499 322-ethoxy-2-[3-(3-fluoroazetidin-1-yl)phenyl]-N-[5- m/z: ES+[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]- [M + H]+1,3,4-thiadiazol-2-yl]acetamide 499 332-ethoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N- m/z: ES+[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]- [M + H]+1,3,4-thiadiazol-2-yl]acetamide 511 342-ethoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]-N- m/z: ES+[5-[[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]amino]- [M + H]+1,3,4-thiadiazol-2-yl]acetamide 511 352-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N- m/z: ES+[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3- [M + H]+yl]amino]-1,3,4-thiadiazol-2-yl]acetamide 499 362-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-N- m/z: ES+[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3- [M + H]+yl]amino]-1,3,4-thiadiazol-2-yl]acetamide 499 372-[3-(3,3-difluoroazetidin-1-yl)-4-fluoro-phenyl]- m/z: ES+2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin- [M + H]+3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide 521 382-[3-(3,3-difluoroazetidin-1-yl)-4-fluoro-phenyl]- m/z: ES+2-methoxy-N-[5-[[(3R)-1-pyridazin-3-ylpyrrolidin- [M + H]+3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide 521 392-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy- m/z: ES+N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin- [M + H]+3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide 517 402-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy- m/z: ES+N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin- [M + H]+3-yl]amino]-1,3,4-thiadiazol-2-yl]acetamide 517 412-[3-(3,3-difluoroazetidin-1-yl)phenyl]-N-[5- m/z: ES+[[(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3- [M + H]+yl]amino]-1,3,4-thiadiazol-2-yl]-2-methoxy- 521 acetamide 42 2-[3-(3,3-difluoroazetidin-l-yl)phenyl]-N-[5- m/z: ES+[[(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3- [M + H]+yl]amino]-1,3,4-thiadiazol-2-yl]-2-methoxy- 521 acetamide 432-(3,5-dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)- m/z: ES+1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4- [M + H]+thiadiazol-2-yl]acetamide 472 442-(3,5-dimethoxyphenyl)-2-methoxy-N-[5-[[(3R)- m/z: ES+1-pyridazin-3-ylpyrrolidin-3-yl]amino]-1,3,4- [M + H]+thiadiazol-2-yl]acetamide 472

Example 45(a) and 45(b)(2S)-3-Methoxy-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-phenyl-propanamideand(2R)-3-Methoxy-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-phenyl-propanamide

N2-[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(Intermediate 6, 0.11 g, 0.389 mmol) and 3-methoxy-2-phenyl-propanoicacid (0.08 g, 0.433 mmol) were dissolved in DMF (2 mL) at r.t under N₂.The mixture was stirred for 5 mins before addition of DIPEA (0.34 mL,1.943 mmol), andN-[(Dimethylamino)(3H-[1,2,3]triazolo[4,5-b]pyridin-3-yloxy)methylidene]-N-methylmethylaminiumhexafluorophosphate, HATU (0.4 mL, 0.361 mmol), then left to sitovernight at room temperature. The crude mixture was passed through a 5g SCX-2 column, washed with MeOH then eluted with 2 M NH₃ in MeOH. Thepure fractions were combined and solvent was removed under reducedpressure and the residue was further purified by preparative HPLC(SunFire C18 column, 5 μm, 50 mm×19 mm, flow rate 25 mL/min).Decreasingly polar ratios of water and MeCN containing 0.1% formic acidwere used as a mobile phase. Pure fractions were combined, evaporatedunder reduced pressure and passed through an SCX-2 column washed withMeOH then eluted with 2 M NH₃ in MeOH. The basic fraction was evaporatedto dryness to afford the mixture of diastereoisomers as a pale yellowsolid. The diastereoisomers were separated by preparative HPLC(Phenomonex Lux C1 column, 20 μm silica, 50 mm diameter, 250 mm length),using a 95/5 mixture of MeCN/MeOH as eluents at 120 mL/min. Fractionscontaining the desired compounds were evaporated to dryness to afford:

First eluted isomer example 45(a)3-Methoxy-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-phenyl-propanamide(31.7 mg, 20%). ¹H NMR (400 MHz, DMSO, 30° C.) 1.95-2.12 (1H, m),2.21-2.35 (1H, m), 2.42 (3H, s), 3.26 (3H, s), 3.4-3.63 (4H, m), 3.74(1H, dd), 3.98 (1H, t), 4.12 (1H, dd), 4.31-4.43 (1H, m), 6.82 (1H, d),7.15-7.46 (6H, m), 7.62 (1H, d), 12.18 (1H, s). m/z: ES⁺ [M+H]⁺ 440.

Second eluted isomer example 45(b)3-Methoxy-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-2-phenyl-propanamide(28.3 mg, 18%).

¹H NMR (400 MHz, DMSO, 30° C.) 1.99-2.12 (1H, m), 2.2-2.36 (1H, m), 2.41(3H, s), 3.26 (3H, s), 3.39-3.62 (4H, m), 3.73 (1H, dd), 3.98 (1H, t),4.12 (1H, dd), 4.31-4.42 (1H, m), 6.82 (1H, d), 7.17-7.46 (5H, m), 7.62(1H, d), 12.19 (1H, s). m/z: ES⁺ [M+H]⁺ 440.

ADDITIONAL EXAMPLES

The compounds of the following Examples were prepared in a similarfashion to the Examples above.

MS data Example m/z: ES⁺ no. Name [M + H]⁺ 46(a) and(2R)-2-ethoxy-2-phenyl-N-(5-{[(3R)-1-(1,2,4- 427 and 46(b)triazin-3-yl)-3 -pyrrolidinyl]amino}-1,3,4- 427thiadiazol-2-yl)acetamide and(2S)-2-ethoxy-2-phenyl-N-(5-{[(3R)-1-(1,2,4- triazin-3-yl)-3-pyrrolidinyl]amino}-1,3,4- thiadiazol-2-yl)acetamide 47(a) and(2S)-2-(4-fluorophenyl)-3-methoxy-N-(5-{[(3R)-1- 444 and 47(b)(3-pyridazinyl)-3-pyrrolidinyl]amino}-1,3,4- 444thiadiazol-2-yl)propanamide and(2R)-2-(4-fluorophenyl)-3-methoxy-N-(5-{[(3R)-1-(3-pyridazinyl)-3-pyrrolidinyl]amino}-1,3,4- thiadiazol-2-yl)propanamide48(a) and (2R)-3-methoxy-2-phenyl-N-(5-{[(3R)-1-(3- 426 and 48(b)pyridazinyl)-3-pyrrolidinyl]amino}-1,3,4-thiadiazol- 4262-yl)propanamide and (2R)-3-methoxy-2-phenyl-N-(5-{[(3R)-1-(3-pyridazinyl)-3-pyrrolidinyl]amino}-1,3,4-thiadiazol- 2-yl)propanamide49(a) and (2R)-3-methoxy-2-phenyl-N-(5-{[(3R)-1-(1,2,4- 427 and 49(b)triazin-3-yl)-3-pyrrolidinyl]amino}-1,3,4-thiadiazol- 4272-yl)propanamide and (2S)-3-methoxy-2-phenyl-N-(5-{[(3R)-1-(1,2,4-triazin-3-yl)-3-pyrrolidinyl]amino}-1,3,4-thiadiazol- 2-yl)propanamide50(a) and (2R)-2-ethoxy-2-(4-fluorophenyl)-N-(5-{[(3R)-1-(3- 444 and50(b) pyridazinyl)-3-pyrrolidinyl]amino}-1,3,4-thiadiazol- 4442-yl)acetamide and (2S)-2-ethoxy-2-(4-fluorophenyl)-N-(5-{[(3R)-1-(3-pyridazinyl)-3-pyrrolidinyl]amino}-1,3,4-thiadiazol- 2-yl)acetamide51(a) and (2S)-2-ethoxy-2-(4-fluorophenyl)-N-(5-{[(3R)-1- 445 and 51(b)(1,2,4-triazin-3-yl)-3-pyrrolidinyl]amino}-1,3,4- 445thiadiazol-2-yl)acetamide and (2R)-2-ethoxy-2-(4-fluorophenyl)-N-(5{[(3R)-1- (1,2,4-triazin-3-yl)-3-pyrrolidinyl]amino}-1,3,4-thiadiazol-2-yl)acetamide 52(a) and(2S)-2-ethoxy-2-(4-fluoro-3-methoxyphenyl)-N-(5- 475 and 52(b){[(3R)-1-(1,2,4-triazin-3-yl)-3-pyrrolidinyl]amino}- 4751,3,4-thiadiazol-2-yl)acetamide and(2R)-2-ethoxy-2-(4-fluoro-3-methoxyphenyl)-N-(5-{[(3R)-1-(1,2,4-triazin-3-yl)-3-pyrrolidinyl]amino}-1,3,4-thiadiazol-2-yl)acetamide 53(a) and(2R)-N-[5-[[(3R)-1-(6-fluoropyridazin-3- 444 and 53(b)yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-3- 444methoxy-2-phenyl-propanamide and(2S)-N-[5-[[(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]-3-methoxy-2-phenyl-propanamide 54(a) and(2R)-3-methoxy-N-[5-[[(3R)-1-(6-methylpyridazin- 440 and 54(b)3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]- 4402-phenyl-propanamide and(2R)-3-methoxy-N-[5-[[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]amino]-1,3,4-thiadiazol-2-yl]- 2-phenyl-propanamide

Intermediate 1N′-[(3R)-1-Pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine

Into a 1 L round-bottom flask was placed a solution of(3R)-1-pyridazin-3-ylpyrrolidin-3-amine dihydrochloride (Intermediate 2,10.5 g, 44.29 mmol) in DMF (400 mL), 5-bromo-1,3,4-thiadiazol-2-amine(7.94 g, 44.10 mmol) and DIPEA (17.07 g, 132.08 mmol). The solution wasstirred for 4 h at 80° C. The resulting mixture was concentrated undervacuum. The crude product was purified by re-crystallization fromethanol/EtOAc to giveN-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamineas a light yellow solid (11 g, 94%). 1H NMR (500 MHz, DMSO-d6, 30° C.) δ2.04 (1H, td), 2.22-2.31 (1H, m), 3.43-3.62 (3H, m), 3.72 (1H, dd), 4.28(1H, dq), 6.27 (2H, s), 6.86 (1H, dd), 7.07 (1H, d), 7.33 (1H, dd), 8.48(1H, dd). m/z: ES⁺ [M+H]⁺ 264.28.

Intermediate 1 was also prepared on a large scale according to thefollowing alternative procedure:

(R)-1-(Pyridazin-3-yl)pyrrolidin-3-amine (Intermediate 3, free baseform, 25.5 g, 150.63 mmol) and 5-bromo-1,3,4-thiadiazol-2-amine (29.8 g,165.70 mmol) with DIPEA (39.4 mL, 225.95 mmol) was agitated as a slurryin MeOH (200 mL) at 45° C. The slurry was cooled to 20° C. and the solidisolated by vacuum filtration. 50 ml MeOH was used as a displacementwash of the filter cake and it was then dried overnight in the vacuumoven at 40° C. Intermediate 1 (32.9 g, 83%) was obtained as a freeflowing beige powder.

Intermediate 2 (3R)-1-Pyridazin-3-ylpyrrolidin-3-amine dihydrochloride

Into a 1 L round-bottom flask was placed a solution of tert-butylN-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]carbamate (Intermediate 4, 20 g,75.66 mmol) in dioxane (200 mL) and concentrated HCl (100 mL). Thesolution was stirred for 30 mins at r.t. The resulting mixture wasconcentrated under vacuum. The crude product was re-crystallized fromMeOH/EtOAc in the ratio of 1:2. This resulted in(3R)-1-pyridazin-3-ylpyrrolidin-3-amine dihydrochloride as an off-whitesolid (13.4 g, 75%). ¹H NMR (300 MHz, DMSO-d6, 26° C.) δ 2.25-2.43 (2H,m), 3.66-3.74 (1H, m), 3.78-3.90 (3H, m), 4.02-4.10 (1H, m), 7.75 (1H,d), 7.94 (1H, dd), 8.66 (1H, d), 8.77-8.98 (3H, brm). m/z: ES⁺ [M+H]⁺165.

Intermediate 3 (free base form) was prepared according to the followingprocedure: tert-butylN-[(3R)-1-(6-Chloropyridazin-3-yl)pyrrolidin-3-yl]carbamate(Intermediate 5, 20 g, 107.38 mmol) in pyridine (400 mL) was mixed withpalladium hydroxide on carbon (Pearlman's Catalyst, 27.5 g, 25.84 mmol)and 1-methyl-1,4-cyclohexadiene (31.0 ml, 276.13 mmol) in MeOH (1375mL). The reaction mixture was then heated to 65° C. for 90 minutes. Withcomplete conversion observed, the reaction was cooled back to ambienttemperature and the catalyst removed by filtration. 3M HCl in MeOH (184mL, 552.27 mmol) was then charged to the reaction mixture, and thesolution heated to 65° C. for 1 h. With complete conversion observed,the reaction solution was cooled back to ambient and passed through10×50 g SCX columns which had been pre-eluted with MeOH. The compoundwas released from the SCX using IM NH₃ in MeOH. The resulting solutionwas diluted with toluene (1 L) and concentrated to dryness via rotaryevaporation to give a free flowing solid.(3R)-1-pyridazin-3-ylpyrrolidin-3-amine was isolated at a strength of97% w/w as the free base.

Intermediate 4 tert-ButylN-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]carbamate

Into a 2 L round-bottom flask was placed a solution of tert-butylN-[(3R)-1-(6-chloropyridazin-3-yl)pyrrolidin-3-yl]carbamate(Intermediate 5, 23 g, 76.98 mmol) in MeOH (800 mL) and Palladium oncarbon (2 g). The system was purged and maintained with Hydrogen gas.The resulting solution was stirred for 4 h at r.t. The solids werefiltered out. The resulting mixture was concentrated under vacuum togive tert-butyl N-[(3R)-1-pyridazin-3-ylpyrrolidin-3-yl]carbamate (20 g,84%) as a yellow solid. ¹H NMR (300 MHz, CDCl₃, 24° C.) δ 1.44 (9H, s),2.25-2.35 (2H, m), 3.48-3.56 (1H, m), 3.70-4.10 (3H, m), 4.35-4.42 (1H,m), 7.26-7.32 (1H, m), 7.70-7.75 (1H, m), 8.53-8.55 (1H, m). m/z: ES⁺[M+H]⁺ 265.

Intermediate 5 tert-ButylN-[(3R)-1-(6-chloropyridazin-3-yl)pyrrolidin-3-yl]carbamate

Into a 1 L round-bottom flask was placed a solution of tert-butylN-[(3R)-pyrrolidin-3-yl]carbamate (20 g, 107.38 mmol) in pyridine (400mL) and 3,6-dichloropyridazine (16 g, 107.40 mmol). The resultingsolution was heated to reflux for overnight. The resulting mixture wasconcentrated under vacuum. The crude product was purified byre-crystallization from ethanol/Et₂O in the ratio of 1:3 to givetert-butyl N-[(3R)-1-(6-chloropyridazin-3-yl)pyrrolidin-3-yl]carbamate(23 g, 72%) as a yellow solid.

¹H NMR (400 MHz, CDCl₃, 30° C.) δ 1.45 (9H, s), 2.02 (1H, dq), 2.31 (1H,td), 3.41 (1H, dd), 3.54-3.70 (2H, m), 3.78 (1H, dd), 4.37 (1H, s), 4.76(1H, s), 6.61 (1H, d), 7.17 (1H, d). m/z: ES⁺ [M+H]⁺ 299.

Intermediate 6N2-[(3R)-1-(6-Methylpyridazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine

5-Bromo-1,3,4-thiadiazol-2-amine (912 mg, 5.07 mmol),(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-amine (Intermediate 7, 860mg, 4.83 mmol) and DIPEA (0.924 mL, 5.31 mmol) were dissolved in DMF (10mL). The reaction was heated to 100° C. for 1 h then left at r.t.overnight. The crude product was purified by ion exchangechromatography, using an SCX column. The desired product was eluted fromthe column using 1M NH₃ in MeOH and pure fractions were evaporated todryness to afford crude product. This was dissolved in DCM/MeOH,adsorbed onto silica and purified by FCC (SiO₂, 0 to 20% MeOH in DCM).Pure fractions were evaporated to dryness to affordN2-[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(350 mg, 26%) as a brown gum. m/z: ES⁺ [M+H]⁺ 278.

Intermediate 7 (3R)-1-(6-Methylpyridazin-3-yl)pyrrolidin-3-amine

Trifluoroacetic acid (12 mL) was added to tert-butylN-[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]carbamate(Intermediate 8, 2.1 g, 7.54 mmol), in DCM (60 mL) at 21° C. undernitrogen. The resulting solution was stirred at 21° C. for 2 h. Thecrude product was purified by ion exchange chromatography, using an SCXcolumn. The desired product was eluted from the column using 7M NH₃ inMeOH and pure fractions were evaporated to dryness to afford(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-amine (1.6 g, 119%) as ayellow oil which solidified on standing. ¹H NMR (400 MHz, DMSO-d6, 27°C.) δ 1.56-1.8 (1H, m), 2.04 (1H, m), 2.39 (3H, s), 3.07 (1H, m),3.37-3.43 (1H, m), 3.47-3.66 (3H, m), 4.08 (1H, s), 6.73 (1H, d), 7.19(1H, d). m/z: ES⁺ [M+H]⁺ 179.

Intermediate 8 tert-ButylN-[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]carbamate

A mixture of DIPEA (8.49 mL, 48.62 mmol), tert-butylN-[(3R)-pyrrolidin-3-yl]carbamate (3.62 g, 19.45 mmol),3-chloro-6-methylpyridazine (2.5 g, 19.45 mmol) and n-butanol (30 mL)was stirred at 130° C. for 12 h then left to cool over the weekend. Thereaction mixture was evaporated and the crude product was purified byFCC (SiO₂, 0 to 10% 1M NH₃ in MeOH in EtOAc). Pure fractions wereevaporated to dryness to afford tert-butylN-[(3R)-1-(6-methylpyridazin-3-yl)pyrrolidin-3-yl]carbamate (2.1 g,38.8%) as a yellow solid. ¹H NMR (400 MHz, DMSO-d6, 27° C.) δ 1.39 (9H,s), 1.88 (1H, m), 2.14 (1H, m), 2.40 (3H, s), 3.23 (1H, m), 3.37-3.45(1H, m), 3.47-3.58 (1H, m), 3.61 (1H, m), 3.99-4.2 (1H, m), 6.77 (1H,d), 7.20 (2H, m). m/z: ES⁺ [M+H]⁺ 279.

Intermediate 9N2-[(3R)-1-(6-Fluoropyridazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine

DIPEA (3.48 mL, 19.96 mmol) was added to5-bromo-1,3,4-thiadiazol-2-amine (1.797 g, 9.98 mmol) and(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3-amine (Intermediate 10, 2 g,10.98 mmol) in anhydrous DMF (40 mL) at r.t. The resulting solution wasstirred at 80° C. for 4 h. The crude product was purified by ionexchange chromatography, using an SCX column. The desired product waseluted from the column using 1M NH₃ in MeOH and pure fractions wereevaporated to dryness to affordN2-[(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine(2.9 g, 103%) as a brown solid. ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ1.90-2.12 (1H, m), 2.23 (1H, dtd), 3.42 (1H, dd), 3.47-3.61 (2H, m),3.69 (1H, dd), 4.25 (1H, dq), 6.25 (2H, s), 7.04 (1H, d), 7.14 (1H, dd),7.33 (1H, dd). m/z: ES⁺ [M+H]⁺ 282.

Intermediate 10 (3R)-1-(6-Fluoropyridazin-3-yl)pyrrolidin-3-amine

tert-Butyl N-[(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3-yl]carbamate(Intermediate 11, 6 g, 21.25 mmol) was added to DCM (70 mL) and TFA(14.00 mL) at 25° C. The resulting solution was stirred at 25° C. for 4h. The crude product was purified by ion exchange chromatography, usingan SCX column. The desired product was eluted from the column using 1MNH₃ in MeOH and pure fractions were evaporated to dryness to afford(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3-amine (2.0 g, 52%) as a paleyellow gummy solid. ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ 1.55-1.83 (1H,m), 1.98-2.13 (1H, m), 2.89-3.14 (1H, m), 3.29-3.43 (1H, m), 3.54 (3H,ddt), 7.06 (1H, dd), 7.30 (1H, dd). m/z. ES⁺ [M+H]⁺ 183.

Intermediate 11 tert-butylN-[(3R)-1-(6-Fluoropyridazin-3-yl)pyrrolidin-3-yl]carbamate

A mixture of 3,6-difluoropyridazine (6.06 g, 52.21 mmol) tert-butylN-[(3R)-pyrrolidin-3-yl]carbamate (9.72 g, 52.21 mmol), DIPEA (22.80 mL,130.53 mmol) and n-butanol (140 mL) was stirred at 130° C. for 10 h. Thereaction mixture was diluted with EtOAc (750 mL), and washed twice withwater (150 mL). The organic layer was dried over Na₂SO₄, filtered andevaporated to afford crude product. This was then dissolved in DCM andthe crude product was purified by FCC (SiO₂, 30-65% EtOAc in heptanes).Pure fractions were evaporated to dryness to afford tert-butylN-[(3R)-1-(6-fluoropyridazin-3-yl)pyrrolidin-3-yl]carbamate (15 g, 102%)as a cream solid. ¹H NMR (400 MHz, CDCl₃, 30° C.) δ 1.46 (9H, s),1.91-2.13 (1H, m), 2.32 (1H, dq), 3.40 (1H, dd), 3.56-3.72 (2H, m), 3.78(1H, dd), 4.37 (1H, s), 4.70 (1H, s), 6.78 (1H, dd), 6.98 (1H, dd). m/z:ES⁺ [M+H]⁺ 283.

Intermediate 12N2-[(3R)-1-(1,2,4-Triazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamine

5-bromo-1,3,4-thiadiazol-2-amine (1.31 g, 7.264 mmol) and(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-amine (Intermediate 13, 1.2 g,7.264 mmol) were combined in DMF (15 mL) at r.t. under N₂. The mixturewas stirred at r.t. overnight. It was then evaporated to dryness andpurified by FCC (SiO₂, 5-10% 2N NH₃ in MeOH in DCM) to giveN2-[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]-1,3,4-thiadiazole-2,5-diamineas a beige foam (1.7 g, 88%). ¹H NMR (400 MHz, DMSO, 30° C.) δ 1.80-1.85(1H, m), 1.98-2.05 (1H, m), 3.40 (4H, brs), 4.03 (1H, brs), 6.09 (2H,s), 6.88 (1H, d), 8.10 (1H, d), 8.38 (1H, d).

Intermediate 13 (3R)-1-(1,2,4-Triazin-3-yl)pyrrolidin-3-amine

tert-Butyl N-[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]carbamate(Intermediate 14, 2.39 g, 9.01 mmol) was dissolved in a mixture of DCM(20 mL) and trifluoroacetic acid (5 mL) and the solution allowed tostand for 1 h at r.t. before being evaporated under reduced pressure.The residue was dissolved in MeOH and passed through a 20 g SCXcartridge flushing with MeOH followed by 3N NH₃ in MeOH to bring off theproduct. The solvent was evaporated under reduced pressure to yield(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-amine (1.460 g, 98%) as a yellowsolid. ¹H NMR (400 MHz, CDCl₃, 27° C.) δ 1.80-1.92 (1H, m), 2.18-2.29(1H, m), 3.45 (1H, s), 3.6-4.01 (4H, m), 8.13 (1H, d), 8.50 (1H, d).m/z: ES⁺ [M+H]⁺ 166.

Intermediate 14 tert-ButylN-[(3R)-1-(1,2,4-Triazin-3-yl)pyrrolidin-3-yl]carbamate

3-Methylsulfanyl-1,2,4-triazine (Intermediate 15, 1.5 g, 11.80 mmol),and tert-butyl N-[(3R)-pyrrolidin-3-yl]carbamate (2.64 g, 14.15 mmol)were dissolved in ethanol (12 mL) and sealed into a microwave tube. Thereaction was heated to 100° C. for 24 h in the microwave reactor andcooled to r.t. LC/MS showed 61% product and 34% unreacted triazine.Further tert-butyl N-[(3R)-pyrrolidin-3-yl]carbamate (0.52 g) was addedand heating at 100° C. in the microwave continued for 15 h. LC/MS showed76% product and 18% unreacted triazine. The solvent was removed underreduced pressure and the residue partitioned between EtOAc and aqueoussodium bicarbonate. The aqueous layer was re-extracted with fresh EtOAcand the combined organics were dried (MgSO₄) filtered and evaporatedunder reduced pressure. The crude product was purified by FCC (SiO₂, 0to 80% EtOAc in heptanes). Relevant fractions were evaporated to givetert-butyl N-[(3R)-1-(1,2,4-triazin-3-yl)pyrrolidin-3-yl]carbamate(2.390 g, 76%) as a yellow solid. ¹H NMR (400 MHz, CDCl₃, 27° C.) δ 1.46(9H, s), 1.96-2.07 (1H, m), 2.26-2.37 (1H, m), 3.55 (1H, s), 3.75 (2H,s), 3.90 (1H, s), 4.39 (1H, s), 4.69 (1H, s), 8.14 (1H, d), 8.53 (1H,d). m/z: ES⁻ [M−H]⁻ 264.

Intermediate 15 3-Methylsulfanyl-1,2,4-triazine

A solution of methyl hydrazinecarbimidothioate hydroiodide (Intermediate16, 7.5 g, 32.18 mmol) in ice/water (400 mL) was added to a stirredsolution of 40% oxalaldehyde (14.70 mL, 128.71 mmol), and sodiumbicarbonate (6.76 g, 80.45 mmol) in ice/water (400 mL) cooled to 0° C.The resulting solution was stirred at 0° C. for 5 h, then extracted withDCM (2×150 mL). The extracts were combined washed with 1M citric acid(50 mL), dried (MgSO₄) and evaporated to give3-methylsulfanyl-1,2,4-triazine (3.60 g, 88%) as a yellow solid. ¹H NMR(400 MHz, CDCl₃, 27° C.) δ 2.68 (3H, s), 8.38 (1H, d), 8.94 (1H, d).

Intermediate 16 Hydrazinecarbimidothioate hydroiodide

Iodomethane (0.623 mL, 10.00 mmol) was added to hydrazinecarbothioamide(0.911 g, 10 mmol), in ethanol (10 mL). The resulting mixture wasstirred at 70° C. for 30 minutes. The reaction was allowed to cool tor.t. The reaction mixture was then filtered through a Nylon filtercup.The resultant solid was then washed with Et₂O and dried under vacuumovernight to give methyl hydrazinecarbimidothioate hydroiodide (1.810 g,78%) as a white solid that was used without further purification.

Intermediate 17[2-[3-(3-Fluoroazetidin-1-yl)phenyl]-2-methoxy-acetyl]oxylithium

Methyl 2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetate(Intermediate 18, 0.26 g, 1.034 mmol) and lithium hydroxide monohydrate(0.07 g, 1.552 mmol) were dissolved in a mixture of MeOH (5 mL) andwater (2 mL). The reaction was stirred for 2 h at r.t. It was thenevaporated and dried in vacuo over the weekend to give[2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetyl]oxylithium as awhite solid (270 mg, 100%). ¹H NMR (400 MHz, DMSO-d6, 27° C.) δ 3.35(3H, s), 4.08-3.91 (2H, m), 4.29 (2H, ddd), 4.42 (1H, s), 5.78-5.56 (1H,m), 6.50 (1H, ddd), 6.68 (1H, t), 6.93 (1H, dt), 7.25 (1H, t). m/z: ES⁺[M+H]⁺ 240.

Intermediate 18 Methyl2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetate

To a mixture of methyl 2-(3-bromophenyl)-2-methoxyacetate (Intermediate19, 500 mg, 1.93 mmol) and 3-fluoroazetidine hydrochloride (215 mg, 1.93mmol) in toluene (20 mL) was added Ruphos palladium(II) phenethylaminechloride (58.3 mg, 0.07 mmol),dicyclohexyl(2′,6′-diisopropoxy-[1,1′-biphenyl]-2-yl)phosphine (29.7 mg,0.06 mmol) and caesium carbonate (2.20 mg, 6.75 mmol). The reaction wassparged with nitrogen for −5 minutes and then heated to 90° C., stirringunder nitrogen overnight. The reaction was cooled to r.t. before beingdiluted with EtOAc and water. The organic layer was dried (MgSO₄),filtered and the solvent evaporated under reduced pressure. Purificationwas by FCC (SiO₂, 0-25% EtOAc in heptanes). Fractions containing productwere evaporated under reduced pressure to yield methyl2-[3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetate (330 mg, 67%) asan oil. ¹H NMR (400 MHz, CDCl₃, 30° C.) δ 3.40 (3H, s), 3.72 (3H, s),3.90-4.02 (2H, m), 4.19 (2H, ddd), 4.71 (1H, s), 5.30-5.51 (1H, m), 6.44(1H, ddd), 6.54-6.56 (1H, m), 6.83 (1H, d), 7.21 (1H, t). m/z: ES⁺[M+H]⁺ 254.

Intermediate 19 Methyl 2-(3-bromophenyl)-2-methoxyacetate

Sodium (0.345 g, 15.00 mmol) was dissolved in dry MeOH (50 mL) under N2and to this solution was added a solution of methyl2-bromo-2-(3-bromophenyl)acetate (Intermediate 20, 4.2 g, 13.64 mmol) indry MeOH (10 mL). The reaction mixture was heated at 40° C. for 2 hoursand then evaporated under reduced pressure. The residue was treated withaqueous ammonium chloride solution, and the mixture was extracted withEtOAc (50 mL×2). The combined organics were dried (MgSO₄), filtered andevaporated under reduced pressure to yield crude product which waspurified by FCC (SiO₂, 0-12% EtOAc in heptanes). Fractions containingproduct were evaporated to dryness to afford methyl2-(3-bromophenyl)-2-methoxyacetate (2.95 g, 83%) as an oil. ¹H NMR (400MHz, CDCl₃, 30° C.) δ 3.42 (3H, s), 3.74 (3H, s), 4.73 (1H, s), 7.24(1H, t), 7.36-7.39 (1H, m), 7.47 (1H, ddd), 7.61 (1H, t). m/z: TOF MSEI⁺ 257.9880.

Intermediate 20 Methyl 2-bromo-2-(3-bromophenyl)acetate

A mixture of methyl 2-(3-bromophenyl)acetate (4 g, 17.46 mmol) and NBS(3.26 g, 18.33 mmol) and(E)-2,2′-(diazene-1,2-diyl)bis(2-methylpropanenitrile) (0.143 g, 0.87mmol) in carbon tetrachloride (50 mL) was heated to reflux for 4 hoursand then cooled to r.t. The solid was filtered off and discarded, thefiltrate evaporated under reduced pressure. Purification was by FCC(SiO₂, 0-5% EtOAc in heptane). Fractions containing product wereevaporated under reduced pressure to yield methyl2-bromo-2-(3-bromophenyl)acetate (4.2 g, 78%) as an oil. ¹H NMR (400MHz, CDCl₃, 30° C.) δ 3.80 (3H, s), 5.28 (1H, s), 7.21-7.26 (1H, m),7.45-7.48 (1H, m), 7.48-7.5 (1H, m), 7.70 (1H, t). m/z: TOF MS EI⁺305.8891.

Intermediate 21[2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-acetyl]oxylithium

Methyl 2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-acetate(Intermediate 23, 0.08 g, 0.295 mmol) and lithium hydroxide monohydrate(0.01 g, 0.354 mmol) were dissolved in a mixture of methanol (3 mL) andwater (2 mL). The reaction was stirred for 2 h at room temperature. Thereaction mixture was evaporated under reduced pressure and dried in thevacuum oven over the weekend to give[2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-acetyl]oxylithium. ¹HNMR (400 MHz, DMSO-d6) δ 3.19 (3H, s), 4.24-4.17 (4H, m), 6.42-6.35 (1H,m), 6.57 (1H, s), 6.82 (1H, d), 7.09 (1H, t).

Intermediate 22 2-[3-(3,3-Difluoroazetidin-1-yl)phenyl]-2-methoxy-aceticacid

A solution of 2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-acetate(Intermediate 23, 270 mg, 1.00 mmol) in MeOH (4 mL) was treated with asolution of lithium hydroxide monohydrate (84 mg, 1.99 mmol) in water (2mL) and the mixture stirred at r.t. for 2 hours. The MeOH was evaporatedunder reduced pressure and the aqueous layer was diluted with aqueousbrine (2 ml), neutralised with acetic acid and extracted with2-methyltetrahydrofuran (4×5 mL). The combined organics were dried(MgSO₄), filtered and evaporated under reduced pressure to yield2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-acetic acid (130 mg,50.8%) as a gum. m/z: ES⁺ [M+H]⁺ 258.

Intermediate 23 Methyl2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-acetate

To a mixture of methyl 2-(3-bromophenyl)-2-methoxyacetate (Intermediate19, 473 mg, 1.83 mmol) and 3,3-difluoroazetidine hydrochloride (236 mg,1.83 mmol) in toluene (20 mL) was added Ruphos palladium (II)phenethylamine chloride (55.2 mg, 0.07 mmol),dicyclohexyl(2′,6′-diisopropoxy-[1,1′-biphenyl]-2-yl)phosphine (28.1 mg,0.06 mmol) and caesium carbonate (2.08 g, 6.39 mmol). The reaction wassparged with N₂ for −5 minutes and then heated to 90° C., with stirringunder N₂ overnight. The reaction was cooled to r.t. before being dilutedwith EtOAc (20 mL) and water (20 mL). The organic layer was dried(MgSO₄), filtered and the solvent evaporated under reduced pressure.Purification was by FCC, (SiO₂, 0-20% EtOAc in heptanes). Fractionscontaining product were evaporated under reduced pressure to yieldmethyl 2-[3-(3,3-difluoroazetidin-1-yl)phenyl]-2-methoxy-acetate (280mg, 57%) as an oil. ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ 3.62 (3H, s),4.25 (4H, t), 4.82 (1H, s), 6.51-6.56 (2H, m), 6.79 (1H, d), 7.19-7.25(1H, m). m/z: ES⁺ [M+H]⁺ 272.

Intermediate 24[2-Methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetyl]oxylithium

Methyl 2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetate(Intermediate 26, 0.24 g, 0.886 mmol) and lithium hydroxide monohydrate(0.06 g, 1.329 mmol) were dissolved in a mixture of methanol (5 mL) andwater (2 mL). The reaction was stirred for 2 h at r.t, then evaporatedunder reduced pressure and dried in vacuo over the weekend. ¹H NMR (400MHz, DMSO-d6) δ 2.96 (3H, s), 3.03 (3H, s), 3.37-3.26 (2H, m), 3.80 (2H,dd), 4.01 (1H, s), 4.09 (1H, tt), 6.06 (1H, dd), 6.25 (1H, t), 6.49 (1H,dt), 6.83 (1H, t).

Intermediate 25 2-Methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]aceticacid

A solution of methyl2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetate (Intermediate 26,450 mg, 1.61 mmol) in MeOH (7 mL) was treated with a solution of lithiumhydroxide monohydrate (135 mg, 3.22 mmol) in water (3 mL) and themixture stirred overnight at r.t. The MeOH was evaporated under reducedpressure and the aqueous residue extracted with ether. The aqueous layerwas acidified with acetic acid, treated with solid sodium chloride togive a saturated solution and extracted with EtOAc (3×10 mL). Theorganic layer was dried (MgSO₄), filtered and evaporated under reducedpressure to yield 2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]aceticacid (236 mg, 58.3%) as a gum. ¹H NMR (400 MHz, DMSO-d6, 30° C.) δ 3.23(3H, s), 3.26 (3H, s), 3.51-3.59 (2H, m), 3.97-4.06 (2H, m), 4.26-4.34(1H, m), 4.59 (1H, s), 6.38 (1H, dd), 6.43 (1H, s), 6.69 (1H, d), 7.13(1H, t). m/z: ES⁺ [M+H]⁺ 252.

Intermediate 26 Methyl2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetate

To a mixture of methyl 2-(3-bromophenyl)-2-methoxyacetate (Intermediate19, 524 mg, 2.02 mmol) and 3-methoxyazetidine hydrochloride (250 mg,2.02 mmol) in toluene (20 mL) was added Ruphos palladium(II)phenethylamine chloride (61.1 mg, 0.07 mmol),dicyclohexyl(2′,6′-diisopropoxy-[1,1′-biphenyl]-2-yl)phosphine (31.2 mg,0.07 mmol) and caesium carbonate (2307 mg, 7.08 mmol). The reaction wassparged with nitrogen for ˜5 minutes and then heated to 90° C., stirringunder nitrogen overnight. The reaction was cooled to r.t. before beingdiluted with EtOAc and water. The organic layer was dried (MgSO₄),filtered and the solvent evaporated under reduced pressure. Purificationby FCC (SiO₂, 0-25% EtOAc in heptanes) gave methyl2-methoxy-2-[3-(3-methoxyazetidin-1-yl)phenyl]acetate (450 mg, 84%) asan oil. ¹H NMR (400 MHz, CDCl₃, 30° C.) δ 3.33 (3H, s), 3.39 (3H, s),3.68-3.74 (5H, m), 4.07-4.13 (2H, m), 4.29-4.36 (1H, m), 4.70 (1H, s),6.43 (1H, ddd), 6.51-6.54 (1H, m), 6.79 (1H, d), 7.19 (1H, t). m/z: ES⁺[M+H]⁺ 266.

Intermediate 27 2-Ethoxy-2-(3-methoxyphenyl)acetic acid

To a stirred mixture of 3-methoxybenzaldehyde (5.0 g, 36.72 mmol) andbromoform (3.85 mL, 44.06 mmol) in ethanol (40 mL) at 0° C. was addeddropwise over a 1 hour period a solution of potassium hydroxide (11.33g, 201.98 mmol) in ethanol (60 mL). After the addition was complete themixture was left to stir at r.t. overnight. A precipitate had formedwhich was removed by filtration, and the filtrate was evaporated to givea paste which was taken up in water (100 mL) and extracted with EtOAc(2×100 mL). The aqueous phase was then acidified to pH=2 with 2M HCl andextracted with EtOAc (2×100 mL). The combined organics were dried(MgSO₄), filtered and evaporated to give a pale brown oil. This wasabsorbed onto silica and was purified by FCC (SiO₂, 5% MeOH in DCM) togive 2-ethoxy-2-(3-methoxyphenyl)acetic acid (3.1 g, 40%) as a palebrown oil. ¹H NMR (400 MHz, CDCl₃, 21° C.) δ 1.28 (3H, t), 2.09 (1H, s),3.64-3.52 (2H, m), 3.81 (3H, s) 4.86 (1H, s), 6.89 (1H, ddd), 6.99 (1H,m), 7.03 (1H, m), 7.29 (1H, t). m/z: ES⁻ [M−H]⁻ 209.

Intermediate 28 2-(3,5-Dimethoxyphenyl)-2-methoxy-acetic acid

A solution of potassium hydroxide (9.79 g, 174.43 mmol) in MeOH (80 mL)was added over 2 h in small portions to a stirred mixture of3,5-dimethoxybenzaldehyde (5.27 g, 31.71 mmol) and bromoform (3.33 mL,38.06 mmol) in MeOH (40 mL) at 0° C. The mixture was then allowed towarm to r.t. and left to stir overnight. Solids were filtered underreduced pressure, and rinsed with MeOH (40 mL). The filtrate wasevaporated to a thick white paste and then re-dissolved in water (150mL). This was washed with Et₂O (200 mL) and the aqueous portionacidified to pH=2 with 2M HCl. The aqueous phase was extracted withEtOAc (500 mL). The combined organics were dried (MgSO₄), filtered andevaporated under reduced pressure to give2-(3,5-dimethoxyphenyl)-2-methoxy-acetic acid as a yellow gum (5.00 g,70%) ¹H NMR (400 MHz, DMSO-d6, 27° C.) δ 3.72 (9H, s), 4.68 (1H, s),6.45 (1H, s), 6.52 (2H, s).

Intermediate 29 3-Methoxy-2-(3-methoxyphenyl)propanoic acid

Ethyl 3-methoxy-2-(3-methoxyphenyl)propanoate (Intermediate 30, 0.3 g,1.259 mmol) was suspended in water (5 mL) and treated with lithiumhydroxide (0.3 g, 12.59 mmol) then heated at 60° C. for 3 hours. Thereaction mixture was allowed to cool to r.t. The aqueous was extractedwith EtOAc (20 mL) and the organics were discarded. The aqueous wasacidified with 1 M HCl to pH=1 and extracted with EtOAc (2×20 mL). Theorganics were dried (MgSO₄), filtered and evaporated under reducedpressure to give a colourless gum which was purified by FCC (SiO₂,0-100% EtOAc in cyclohexane). Fractions containing the desired compoundwere combined and evaporated to give3-methoxy-2-(3-methoxyphenyl)propanoic acid (88 mg, 33%). ¹H NMR (400MHz, DMSO-d6, 30° C.) δ 3.25 (s, 3H), 3.49 (1H, dd), 3.74 (3H, s),3.88-3.76 (2H, m), 6.93-6.80 (3H, m), 7.32-7.17 (1H, m), 12.51 (1H, s).m/z: ES⁺ [M+H]⁺ 211.

Intermediate 30 Ethyl 3-methoxy-2-(3-methoxyphenyl)propanoate

To a solution of ethyl 3-hydroxy-2-(3-methoxyphenyl)propanoate(Intermediate 31, 0.38 g, 1.695 mmol) in MeCN (2 mL) in a round bottomedflask was added silver oxide (0.5 g, 2.14 mmol). The solution was cooledto 0° C. and treated with iodomethane (0.17 mL, 2.676 mmol). Thereaction mixture was allowed to stir at room temperature in the dark for2.5 days. LCMS analysis showed some product but mainly startingmaterial. A further 0.53 mL of iodomethane and 0.15 g of silver oxidewere added and stirring was continued for 24 hours. The reaction hadprogressed further so it was left to stir for another 7 days. LCMSanalysis showed complete conversion. The mixture was filtered throughcelite and the solvent was removed under reduced pressure. The residuewas adsorbed onto silica and purified by FCC (SiO₂, 0-40% EtOAc incyclohexane). The appropriate fractions were evaporated under reducedpressure to isolate ethyl 3-methoxy-2-(3-methoxyphenyl)propanoate as acolourless oil (0.3 g, 74%). ¹H NMR (400 MHz, CDCl₃, 21° C.) δ 1.23 (3H,t), 3.37 (3H, s), 3.58 (1H, dd), 3.80 (3H, s), 3.84 (1H, dd), 3.97 (1H,t), 4.24-4.08 (2H, m), 6.82 (1H, ddd), 6.95-6.85 (2H, m), 7.23 (1H, d).m/z: ES⁺ [M+H]⁺ 239.

Intermediate 31 Ethyl 3-hydroxy-2-(3-methoxyphenyl)propanoate

Ethyl 2-(3-methoxyphenyl)acetate (1.0 g, 5.149 mmol) was weighed into around bottom flask with sodium hydrogen carbonate (0.02 g, 0.257 mmol).DMSO (8 mL) was added followed by paraformaldehyde (0.39 mL, 5.149mmol). The resultant suspension was allowed to stir at room temperaturefor 24 hours then the reaction was heated at 60° C. for 3 hours to givea colourless solution. The reaction mixture was cooled, diluted withwater (100 mL) and neutralised with 0.5 M HCl. The aqueous was extractedwith EtOAc (3×50 mL), dried (MgSO₄) and evaporated under reducedpressure to give a colourless oil which was purified by FCC (SiO₂, 0-50%EtOAc in cyclohexane). The solvent was removed under reduced pressure toprovide ethyl 3-hydroxy-2-(3-methoxyphenyl)propanoate as a colourlessoil (0.38 g, 33%). ¹H NMR (400 MHz, CDCl₃, 21° C.) δ 1.23 (3H, t), 2.26(1H, s), 3.86-3.76 (5H, m), 4.28-4.06 (3H, m), 6.89-6.79 (3H, m),7.30-7.22 (1H, m). m/z: ES⁺ [M+H]⁺ 225.

Intermediate 32 2-Methoxy-2-[3-(trifluoromethoxy)phenyl]acetic acid

A solution of potassium hydroxide (1.851 g, 33.00 mmol) in MeOH (10 mL)was added over 2 h in small portions to a stirred mixture of3-(trifluoromethoxy)benzaldehyde (1.141 g, 6 mmol) and bromoform (0.630mL, 7.20 mmol) in MeOH (5.00 mL) at 0° C. The mixture was then allowedto warm to r.t. and left to stir overnight. A white precipitate formedin the reaction mixture. The solids were filtered off under reducedpressure rinsing the filter cake with MeOH (15 mL). The filtrate wasevaporated to a thick white paste then re-dissolved in water (50 mL).This was then washed with Et₂O (50 mL). The aqueous phase was acidifiedto pH=2 (˜5 mL 2M HCl solution) and then extracted into EtOAc (3×50 mL).The combined organics were dried (MgSO₄), filtered and evaporated underreduced pressure to give a clear oil. The crude product was purified byFCC (SiO₂, 10-50% EtOAc in heptanes). Pure fractions were evaporated todryness to afford 2-methoxy-2-[3-(trifluoromethoxy)phenyl]acetic acid(0.832 g, 55%) as a colourless oil. ¹H NMR (400 MHz, CDCl₃, 30° C.) δ3.47 (3H, s), 4.81 (1H, s), 7.20-7.24 (1H, m), 7.33 (1H, s), 7.37-7.46(2H, m). m/z: ES⁻ [M−H]⁻ 249.4.

Intermediate 33 2-(3,5-Dimethoxyphenyl)-2-ethoxy-acetic acid

To a stirred mixture of 3,5-dimethoxybenzaldehyde (2.16 mL, 15.04 mmol)and bromoform (1.58 mL, 18.054 mmol) in ethanol (15 mL) at 0° C. wasadded, dropwise over 30 mins, a solution of potassium hydroxide (4.64 g,82.747 mmol) in ethanol (30 mL). After completion of addition themixture was left to stir and warmed to room temperature overnight. Nextmorning, the precipitate was removed by filtration. The filtrate wasevaporated to give a paste which was taken up in water (75 mL) andextracted with EtOAc (2×75 mL). The aqueous phase was then acidified topH=2 with 2N HCl. It was extracted with EtOAc (2×100 mL). The combinedorganics were dried (MgSO₄) and evaporated to give the crude product.Purification was by FCC (SiO₂, 0-5% MeOH in DCM). Pure fractions werecombined and evaporated under reduced pressure to give2-(3,5-dimethoxyphenyl)-2-ethoxy-acetic acid as an orange gum (2.46 g,68%). ¹H NMR (400 MHz, DMSO-d6, 25° C.) δ 1.15 (3H, t), 3.37-3.42 (1H,m), 3.48-3.55 (1H, m), 3.73 (6H, s), 4.78 (1H, s), 6.29 (1H, s), 6.53(1H, s), 6.54 (1H, s), 12.76 (1H, s). m/z: ES⁻ [M−H]⁻ 239.

Intermediate 342-[4-Fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-acetic acid

Methyl 2-[4-fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-acetate(Intermediate 35, 250 mg, 0.88 mmol) was suspended in water (1.5 mL),treated with lithium hydroxide (42 mg, 1.77 mmol) and heated at 40° C.for 2 h. The reaction mixture was allowed to cool to room temperatureand the MeOH evaporated under reduced pressure. The aqueous phase wasdiluted with brine (5 mL) and adjusted to pH=5 by the addition ofsaturated aqueous citric acid. The aqueous was extracted with DCMcontaining 5% MeOH (3×10 mL). The combined organic extracts were dried(MgSO₄) and filtered. The aqueous was further acidified to pH=3 by theaddition of formic acid then re-extracted with EtOAc (4×10 mL). TheEtOAc extracts were combined, dried (MgSO₄), filtered, combined with theDCM/MeOH extracts then evaporated under reduced pressure. The gummyresidue was concentrated twice from Et₂O to give2-[4-fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-acetic acid (221mg, 93%) as a pale yellow glassy solid. ¹H NMR (400 MHz, CDCl₃, 21° C.)δ 3.33 (3H, s), 3.41 (3H, s), 3.80-3.83 (2H, m), 4.19-4.23 (2H, m),4.29-4.33 (1H, m), 4.68 (1H, s), 6.50 (1H, dd), 6.75 (1H, ddd), 6.94(1H, dd). m/z: ES⁺ [M+H]⁺ 270.1.

Intermediate 35 Methyl2-[4-fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-acetate

Methyl 2-(3-bromo-4-fluoro-phenyl)-2-methoxy-acetate (Intermediate 36,600 mg, 2.17 mmol) was weighed into a 3 neck round bottomed flask fittedwith a suba seal, N₂ inlet and reflux condenser. The flask was flushedwith N₂ for 5 minutes then 3-methoxyazetidine hydrochloride (0.27 g,2.17 mmol) and caesium carbonate (2.47 g, 7.58 mmol) were added. In aseparate flask 2-dicyclohexylphosphino-2,6-di-iso-propoxybiphenyl,(Ruphos, 0.04 g, 0.087 mmol) and Ruphos Pd G2 (0.06 g, 0.077 mmol) weredissolved in dry toluene. The solution was degassed with N₂ for 5minutes through a needle and then added to the other reagents in the 3neck round bottom flask through the suba seal. The reaction mixture washeated at 90° C. for 20 h. The reaction mixture was allowed to cool toroom temperature and diluted with EtOAc (20 mL) and water (20 mL). Theorganic layer was collected and the aqueous was washed with a furtherportion of EtOAc (20 mL). The organics were washed with brine (50 mL),separated and evaporated under reduced pressure. The residue waspurified by FCC (SiO₂, 0-70% EtOAc in petroleum ether). Pure fractionswere combined and evaporated under reduced pressure to give methyl2-[4-fluoro-3-(3-methoxyazetidin-1-yl)phenyl]-2-methoxy-acetate (136 mg,22%) as a colourless oil. ¹H NMR (400 MHz, CDCl₃, 21° C.) δ 3.33 (3H,s), 3.38 (3H, s), 3.72 (3H, s), 3.78-3.83 (2H, m), 4.19-4.23 (2H, m),4.29-4.33 (1H, m), 4.67 (1H, s), 6.55 (1H, dd), 6.75 (1H, ddd), 6.92(1H, dd). m/z: ES⁺ [M+H]⁺ 284.0.

Intermediate 36 Methyl 2-(3-bromo-4-fluoro-phenyl)-2-methoxy-acetate

Sodium (93 mg, 4.05 mmol) was dissolved in MeOH (12 mL) under N₂. Asolution of methyl 2-bromo-2-(3-bromo-4-fluoro-phenyl)acetate(Intermediate 37, 1.20 g, 3.68 mmol) in MeOH (3 mL) was added and thereaction mixture heated to 40° C. for 2 h. The reaction mixture wascooled to r.t. and evaporated under reduced pressure. The residue wastreated with saturated NH₄Cl(aq) (10 mL) and extracted with EtOAc (3×10mL). The combined organic extracts were dried (MgSO₄), filtered andevaporated under reduced pressure. The residue was purified by FCC(SiO₂, 0-10% EtOAc in petroleum ether). Pure fractions were combined andevaporated under reduced pressure to give methyl2-(3-bromo-4-fluoro-phenyl)-2-methoxy-acetate (616 mg, 60%) as acolourless oil. ¹H NMR (400 MHz, CDCl₃, 21° C.) δ 3.42 (3H, s), 3.74(3H, s), 4.73 (1H, s), 7.12 (1H, t), 7.37 (1H, ddd), 7.67 (1H, dd).

Intermediate 37 Methyl 2-bromo-2-(3-bromo-4-fluoro-phenyl)acetate

A solution of methyl 2-(3-bromo-4-fluoro-phenyl)acetate (5.0 g, 20.24mmol) in carbon tetrachloride (50 mL) was prepared andN-bromosuccinimide (3.78 g, 21.25 mmol) was added followed by2,2′-azobis(2-methylpropionitrile) (0.17 g, 1.01 mmol). The reaction washeated to reflux for 4 hours and allowed to cool to room temperature.The precipitate was filtered off and the solution was evaporated underreduced pressure. The crude residue was purified by FCC (SiO₂, 0-10%EtOAc in petroleum ether). Pure fractions were combined and evaporatedunder reduced pressure to give methyl2-bromo-2-(3-bromo-4-fluoro-phenyl)acetate (5.30 g, 80%) as a colourlessoil. ¹H NMR (400 MHz, CDCl₃, 21° C.) δ 3.81 (3H, s), 5.28 (1H, s), 7.12(1H, t), 7.49 (1H, ddd), 7.78 (1H, dd).

Intermediate 38 Lithium2-[4-fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetate

Lithium hydroxide (99.86 mg, 4.16 mmol) was suspended in water (10 mL)and methanol (33 mL), treated with methyl2-[4-fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetate(Intermediate 39, 565 mg, 2.08 mmol) and heated at 45° C. under nitrogenfor 2 hours. The reaction mixture was allowed to cool to roomtemperature and the MeOH and water were removed under reduced pressurebefore being dried in a vacuum oven for 1 day. This gave lithium2-[4-fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetate (0.60 g,75%). ¹H NMR (400 MHz, DMSO-d6) δ 3.18 (3H, s), 3.95-3.50 (3H, m),4.18-4.11 (1H, m), 4.21 (1H, s), 5.58-5.32 (1H, m), 6.60 (1H, dd),6.77-6.70 (1H, m), 6.96-6.88 (1H, m). m/z: ES⁺ [M+H]⁺ 263.1.

Intermediate 39 Methyl2-[4-fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetate

Into a 2 neck round bottomed flask fitted with a suba seal, magneticstirrer bar nitrogen inlet and reflux condenser was added Ruphos Pd G1methyl-t-butylether adduct (0.16 g, 0.193 mmol),2-dicyclohexylphosphino-2,6-diisopropoxybiphenyl, (Ruphos, 0.09 g, 0.193mmol), 3-fluoroazetidine hydrochloride (0.5 g, 4.461 mmol) and caesiumcarbonate (4.24 g, 13.01 mmol) the flask was flushed with nitrogen gasfor 5 minutes then in a separate round bottomed flask was placed methyl2-(3-bromo-4-fluoro-phenyl)-2-methoxy-acetate (Intermediate 36, 1.03 g,3.717 mmol). The flask was fitted with a suba seal and toluene (25 mL)was added under nitrogen. The resultant solution was degassed bybubbling nitrogen through it for 5 minutes then it was added undernitrogen to the 2 neck round bottomed flask containing the otherreagents. The reaction mixture was heated at 90° C. for 48 hours andthen allowed to cool to room temperature and evaporated under reducedpressure. The residue was partitioned between EtOAc (100 mL) and water(100 mL). The aqueous layer was extracted with a further portion ofEtOAc (200 mL). The organics were combined, dried with MgSO₄, filteredand evaporated under reduced pressure to yield a light brown oil whichwas dissolved in EtOAc and absorbed onto silica, then purified by flashcolumn chromatography, eluting with 0-100% EtOAc in cyclohexane.Evaporation of the appropriate fractions provided methyl2-[4-fluoro-3-(3-fluoroazetidin-1-yl)phenyl]-2-methoxy-acetate, (565 mg,33%) as a pale yellow liquid. 1H NMR (400 MHz, DMSO-d6) δ 3.28 (3H, s),3.63 (3H, s), 4.02-3.84 (2H, m), 4.31-4.17 (2H, m), 4.84 (1H, s),5.57-5.31 (1H, m), 6.62-6.53 (1H, m), 6.79-6.71 (1H, m), 7.07 (1H, dd).m/z: ES⁺ [M+H]⁺ 271.2.

Intermediate 40 Lithium2-[3-(3-fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-acetate

Lithium hydroxide (0.02 g, 0.932 mmol) was suspended in water (1 mL) andMeOH (3 mL) and treated with methyl2-[3-(3-fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-acetate(Intermediate 41, 0.13 g, 0.466 mmol), The mixture was heated to 45° C.under N₂ for 2 h. It was then allowed to cool to r.t. and the solventsremoved under reduced pressure before being dried in vacuo for 3 days toyield lithium2-[3-(3-fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-acetate (137mg, 101%). m/z: ES⁺ [M+H]⁺ 269.3.

Intermediate 41 Methyl2-[3-(3-fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-acetate

In a round bottomed flask was placed methyl2-(3-bromo-5-methoxy-phenyl)-2-methoxy-acetate (Intermediate 42, 0.3 g,1.038 mmol). The flask was fitted with a suba seal and toluene (6.5 mL)was added under N₂. The resultant solution was degassed by bubbling N₂through for 5 minutes. Into a 2 neck round bottomed flask fitted with asuba seal, magnetic stirrer bar, nitrogen inlet and reflux condenser wasadded Ruphos Pd G1 methyl-t-butylether adduct (0.04 g, 0.054 mmol),2-dicyclohexylphosphino-2,6-diisopropoxybiphenyl, (Ruphos, 0.03 g, 0.054mmol), 3-fluoroazetidine hydrochloride (0.14 g, 1.245 mmol) and caesiumcarbonate (1.18 g, 3.632 mmol). The flask was flushed with N₂ for 5 minbefore addition of the degassed solution of methyl2-(3-bromo-5-methoxy-phenyl)-2-methoxy-acetate (0.3 g, 1.038 mmol) intoluene through the suba seal. The reaction mixture was heated at 90° C.for 48 hours and then allowed to cool to room temperature and evaporatedunder reduced pressure. The residue was partitioned between EtOAc (100mL) and water (100 mL). The aqueous layer was extracted with a furtherportion of EtOAc (200 mL). The organics were combined, dried (MgSO₄),filtered and evaporated to yield a light brown oil which was dissolvedin EtOAc and absorbed onto silica. It was purified by FCC (SiO₂, 0-50%EtOAc in cyclohexane). Evaporation of the appropriate fractions providedmethyl 2-[3-(3-fluoroazetidin-1-yl)-5-methoxy-phenyl]-2-methoxy-acetate(132 mg, 42%) as a pale yellow liquid. ¹H NMR (400 MHz, DMSO-d6, 30° C.)δ 3.27 (3H, s), 3.62 (3H, s), 3.71 (3H, s), 3.78-3.96 (2H, m), 4.05-4.21(2H, m), 4.77 (1H, s), 5.35-5.57 (1H, m), 5.98 (1H, s), 6.05 (1H, s),6.29 (1H, s). m/z: ES⁺ [M+H]⁺ 283.3.

Intermediate 42 Methyl 2-(3-bromo-5-methoxy-phenyl)-2-methoxy-acetate

2-(3-Bromo-5-methoxy-phenyl)-2-methoxy-acetic acid (Intermediate 43,2.77 g, 10.06 mmol) was dissolved in MeOH (30 mL), treated withconcentrated sulfuric acid (0.17 mL, 2.014 mmol), and then heated to 65°C. for 2 h. The mixture was cooled to r.t. evaporated then diluted withwater (5 mL) followed by saturated aqueous sodium bicarbonate (20 mL).The aqueous was extracted into DCM followed by EtOAc. The combinedorganics were evaporated and the crude material purified by FCC (SiO₂,0-40% EtOAc in cyclohexane). Fractions containing the product wereevaporated to give methyl 2-(3-bromo-5-methoxy-phenyl)-2-methoxy-acetate(1.2 g, 41%). (400 MHz, DMSO-d6, 30° C.) δ 3.31 (s, 3H), 3.65 (3H, s),3.78 (3H, s), 4.94 (1H, s), 6.90-6.95 (1H, m), 7.11-7.14 (1H, m), 7.15(1H, dd). m/z: ES⁺ [M+H]⁺ 290.

Intermediate 43 2-(3-Bromo-5-methoxy-phenyl)-2-methoxy-acetic acid

To a stirred mixture of potassium hydroxide (2.87 g, 51.15 mmol) andbromoform (0.98 mL, 11.16 mmol) in methanol (15 mL) at 0° C. was added,over a 10 mins period a suspension of 3-bromo-5-methoxy-benzaldehyde(0.75 mL, 9.301 mmol) in methanol (60 mL). After addition the mixtureleft to stir as it warmed to room temperature overnight. The reactionmixture was then treated with DCM (60 mL) and the precipitate wasremoved by filtration. The filtrate was evaporated to give a paste whichwas taken up in water (200 mL) and extracted with DCM (200 mL). Theaqueous phase was then acidified to pH 2 with 2M HCl and extracted withDCM (2×100 mL) and EtOAc (2×100 mL). The combined organics wereevaporated under reduced pressure and chromatographed (SiO₂, 0-30%EtOAc-cyclohexane). The fractions containing the product were evaporatedto give 2-(3-bromo-5-methoxy-phenyl)-2-methoxy-acetic acid (0.84 g,33%). ¹H NMR (400 MHz, DMSO-d6) δ 3.30 (s, 3H) 3.77 (s, 3H), 4.78 (s,1H), 6.94-6.91 (m, 1H), 7.14-7.10 (m, 2H), 13.04 (s, 1H). m/z: ES⁺[M+H]⁺ 275.1.

Intermediate 44 2-Methoxy-2-[3-methoxy-5-(trifluoromethoxy)phenyl]aceticacid

To a stirred mixture of 3-methoxy-5-(trifluoromethoxy)benzaldehyde(Intermediate 45, 0.15 g, 0.681 mmol) and bromoform (0.07 mL, 0.81 mmol)in MeOH (1 mL) at 0° C. was added, dropwise over 30 mins, a solution ofpotassium hydroxide (0.207 g, 3.75 mmol) in MeOH (2 mL). The mixture wasstirred and warmed to r.t. overnight. The resulting precipitate wasremoved by filtration. The filtrate was evaporated to give a paste whichwas taken up in water (100 mL) and extracted with EtOAc (2×100 mL) toremove unreacted aldehyde. The aqueous phase was then acidified to pH=2with 2N hydrochloric acid and extracted with EtOAc (2×100 mL). Thecombined organics were dried (MgSO₄), filtered and evaporated underreduced pressure to afford crude product which was purified by FCC(SiO₂, 0-10% MeOH in DCM) to give2-methoxy-2-[3-methoxy-5-(trifluoromethoxy)phenyl]acetic acid (191 mg,34%) as a yellow solid. m/z: ES⁺ [M+H]⁺ 280.

Intermediate 45 3-Methoxy-5-(trifluoromethoxy)benzaldehyde

To a stirred solution of [3-methoxy-5-(trifluoromethoxy)phenyl]methanol(Intermediate 46, 330 mg, 1.49 mmol) in DCM (18 mL) was added activatedmanganese dioxide (646 mg, 7.43 mmol) and the reaction mixture wasstirred at r.t. for 3 hours. The reaction mixture was diluted with water(50 mL), extracted with DCM (3×50 mL), washed with brine (100 mL), dried(MgSO₄), filtered and evaporated under reduced pressure to afford crudeproduct which was purified by FCC (SiO₂, 0-50% EtOAc in cyclohexane) togive 3-methoxy-5-(trifluoromethoxy)benzaldehyde, (0.15 g, 45%) as acolourless oil. ¹H NMR (400 MHz, DMSO-d6, 21° C.) 63.83 (3H, s),7.24-7.27 (1H, m), 7.39-7.42 (1H, m), 7.46-7.48 (1H, m), 9.94 (1H, s).m/z: ES⁺ [M+H]⁺ 222.

Intermediate 46 [3-Methoxy-5-(trifluoromethoxy)phenyl]methanol

To a stirred solution of 3-methoxy-5-(trifluoromethoxy)benzoic acid(0.50 g, 2.12 mmol) in THE (10 mL) at 0° C. under nitrogen was added,dropwise over 10 mins, lithium aluminium hydride (1M in THF, 2.33 mL,2.33 mmol). The reaction mixture was stirred and warmed to r.t. undernitrogen for 3 hours. The reaction mixture was diluted with water (10mL), followed by of 2N NaOH (30 mL) solution and the salts formed wereremoved by filtration. The filtrate was extracted with DCM (3×100 mL),dried (MgSO₄), filtered and evaporated under reduced pressure to affordcrude product which was purified by FCC (SiO₂, 0-50% EtOAc incyclohexane) to give [3-methoxy-5-(trifluoromethoxy)phenyl]methanol(0.33 g, 67%) as a white solid. m/z: ES⁺ [M+H]⁺ 222.

Intermediate 47 2-[3-(Difluoromethoxy)phenyl]-2-ethoxy-acetic acid

To a stirred mixture of 3-(difluoromethoxy)benzaldehyde (2.0 g, 11.619mmol) and bromoform (1.22 mL, 13.94 mmol) in ethanol (40 mL) at 0° C.was added, dropwise over 1 hour, a solution of potassium hydroxide (3.59g, 63.90 mmol) in ethanol (20 mL). After addition the mixture was leftto stir and warmed to r.t. overnight. The precipitate was removed byfiltration and the filtrate evaporated under reduced pressure to give apaste which was taken up in water (100 mL) and extracted with EtOAc(2×75 mL). The aqueous phase was then acidified to pH=1 with 2M HCl andextracted with EtOAc (2×75 mL). The combined organics were dried(MgSO₄), filtered and evaporated to give a pale brown oil which waspurified by FCC (SiO₂, 95:5 cyclohexane: EtOAc+0.1% formic acidincreasing to 8:2 EtOAc:cyclohexane+0.1% formic acid). Appropriatefractions were evaporated under reduced pressure to provide2-[3-(difluoromethoxy)phenyl]-2-ethoxy-acetic acid as

a colourless oil (1.8 g, 62%). ¹H NMR (400 MHz, CDCl₃, 21° C.) δ 1.29(3H, t), 3.49-3.70 (2H, m), 4.89 (1H, s), 6.52 (1H, t), 7.07-7.15 (1H,m), 7.21-7.26 (1H, m), 7.28-7.35 (1H, m), 7.34-7.41 (1H, m). m/z: ES⁻[M−H]⁻ 245.

Intermediate 48 2-Methoxy-2-[3-(oxetan-3-yl)phenyl]acetic acid

A solution of potassium hydroxide (1.427 g, 25.43 mmol) in MeOH (30 mL)was added over 2 hours in small portions to a stirred mixture of3-(oxetan-3-yl)benzaldehyde (Intermediate 49, 750 mg, 4.62 mmol) andbromoform (0.485 mL, 5.55 mmol) in MeOH (15 mL) at 0° C. The mixture wasthen allowed to warm to room temperature and left to stir overnight. Thesolids were filtered under reduced pressure, rinsing the solids withMeOH (15 mL). The filtrate was evaporated to a thick white paste thenre-dissolved in water (150 mL). This was washed with Et2O (200 mL) andthen acidified to pH=2 with 2M. The aqueous phase was extracted intoEtOAc (300 mL) and the combined organics were dried (MgSO₄), filteredand evaporated under reduced pressure to give2-methoxy-2-[3-(oxetan-3-yl)phenyl]acetic acid as a colourless oil (1.0g, 97%) which was used without further purification. ¹H NMR (400 MHz,DMSO, 30° C.) 63.31 (3H, s), 4.18-4.35 (1H, m), 4.58 (2H, m), 4.77 (1H,s), 4.94 (1H, m), 7.22-7.57 (4H, m).

Intermediate 49 3-(Oxetan-3-yl)benzaldehyde

Manganese (IV) oxide (14.30 g, 164.43 mmol) was added to(3-(oxetan-3-yl)phenyl)methanol (Intermediate 50, 1.5 g, 8.22 mmol) inDCM (70 mL) at 21° C. under nitrogen. The resulting mixture was stirredat 21° C. for 16 hours. The reaction was then filtered through celite,washed with DCM and evaporated to a gum. The crude product was purifiedby FCC (SiO₂, 10-60% EtOAc in heptanes). Pure fractions were evaporatedto dryness to afford 3-(oxetan-3-yl)benzaldehyde (800 mg, 60%) as acolourless gum. ¹H NMR (400 MHz, DMSO, 30° C.) 64.21-4.51 (1H, m), 4.64(2H, m), 4.98 (2H, m), 7.60 (1H, t), 7.74 (1H, d), 7.79-7.85 (1H, d),7.94 (1H, s), 10.03 (1H, s).

Intermediate 50 (3-(Oxetan-3-yl)phenyl)methanol

Lithium aluminium hydride (1M in THF, 1.94 mL, 1.95 mmol) was addeddropwise to sodium 3-(oxetan-3-yl)benzoate (300 mg, 1.50 mmol) in THE(10 mL) at 0° C. under nitrogen. The resulting mixture was stirred at 0°C. for 20 minutes, then at 25° C. for 18 hours. 5% aqueous HCl was addedand the mixture extracted with EtOAc (100 mL), washed with 2M aqueousNa₂CO₃ solution, before the organic layer was dried (Na₂SO₄), filteredand evaporated. The crude product was purified by FCC (SiO₂, 50-100%EtOAc in heptane followed by 10% 1M NH₃ in MeOH in EtOAc). Purefractions were evaporated to dryness to afford(3-(oxetan-3-yl)phenyl)methanol (140 mg, 57%) as a colourless gum. ¹HNMR (400 MHz, DMSO, 30° C.) δ 4.20 (1H, m), 4.51 (2H, d), 4.62 (2H, m),4.95 (2H, m), 5.16 (1H, t), 7.17-7.29 (2H, m), 7.31-7.41 (2H, m).

Intermediate 51 2-Ethoxy-2-[3-(trifluoromethoxy)phenyl]acetic acid

To a stirred mixture of potassium hydroxide (1.62 g, 28.93 mmol) andbromoform (0.55 mL, 6.31 mmol) in ethanol (15 mL) at 0° C. was added,slowly over a 10 min period, a solution of3-(trifluoromethoxy)benzaldehyde (0.75 mL, 5.26 mmol) in ethanol (30mL). After addition the mixture was left to stir and warmed to roomtemperature overnight. The precipitate was removed by filtration. Thefiltrate was evaporated to give a paste which was taken up in water (200mL) and extracted with DCM (100 mL). This formed an emulsion, theaqueous phase was then acidified with 2M HCl (10 mL) and separated. Itwas then further extracted with EtOAc (100 mL). The combined organicswere evaporated under reduced pressure and purified by FCC (SiO₂, 0-50%EtOAc in cyclohexane). Pure fractions were combined and evaporated underreduced pressure to give 2-ethoxy-2-[3-(trifluoromethoxy)phenyl]aceticacid (690 mg, 49%). ¹H NMR (400 MHz, DMSO-d6, 21° C.) δ 1.16 (3H, t),3.38-3.50 (1H, m), 3.53-3.66 (1H, m), 4.99 (1H, s), 7.28-7.39 (2H, m),7.45 (1H, d), 7.53 (1H, t), 13.04 (1H, s). m/z: ES⁺ [M+H]⁺ 265.

Intermediate 52 2-[4-Fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-aceticacid

Lithium hydroxide (0.02 g, 0.9 mmol) was suspended in water (1 mL) andmethanol (3 mL) and treated with ethyl2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-acetate (Intermediate53, 0.13 g, 0.45 mmol) and heated at 45° C. for 2 hours. The reactionmixture was allowed to cool to room temperature and the MeOH removedunder reduced pressure. The aqueous phase was diluted with brine (20 mL)and adjusted to pH 3 by the addition of formic acid then extracted withEtOAc (3×20 mL). The combined EtOAc extracts were washed with brine (30mL) dried over MgSO₄, filtered and the solvent was removed under reducedpressure to yield2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-acetic acid (0.127 g,105%). ¹H NMR (400 MHz, CDCl₃-d6, 25° C.) δ 3.47 (3H, s), 4.78 (1H, s),7.23 (1H, dd), 7.37-7.46 (2H, m). m/z: ES⁺ [M+H]⁺ 268.98.

Intermediate 53 Ethyl2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-acetate

In a round bottomed flask under N₂ was added sodium (0.04 g, 1.66 mmol)to dry methanol (12 mL). To this solution was added methyl2-bromo-2-[4-fluoro-3-(trifluoromethoxy)phenyl]acetate (Intermediate 54,0.5 g, 1.51 mmol) in methanol (3 mL) under N₂. The reaction mixture washeated at 40° C. for 2 hours. The solvent was removed under reducedpressure and the remaining gum was partitioned between saturatedammonium chloride solution (50 mL) and EtOAc (50 mL). The aqueous layerwas extracted with a further portion of EtOAc (50 mL). The organics werecombined dried with MgSO₄, filtered and the solvent was removed underreduced pressure. The residual oil was purified by flash columnchromatography eluting with 100% cyclohexane to 15% EtOAc incyclohexane. Evaporation of the appropriate fractions provided ethyl2-[4-fluoro-3-(trifluoromethoxy)phenyl]-2-methoxy-acetate (0.127 g,28%). ¹H NMR (400 MHz, CDCl₃-d6, 25° C.) δ 1.23 (3H, t), 3.44 (3H, s),4.12-4.28 (2H, m), 4.74 (1H, d), 7.20 (1H, dd), 7.39 (1H, dddd),7.42-7.48 (m, 1H).

Intermediate 54 Methyl2-bromo-2-[4-fluoro-3-(trifluoromethoxy)phenyl]acetate

Methyl 2-[4-fluoro-3-(trifluoromethoxy)phenyl]acetate (Intermediate 55,0.65 g, 2.578 mmol) and N-bromosuccinimide (4.08 g, 22.919 mmol) wereweighed into a round bottomed flask and2,2-azobis(2-methylpropionitrile), (AIBN, 0.02 g, 0.129 mmol) in carbontetrachloride (6 mL) were added. The reaction was heated to reflux for 4hours and allowed to cool to room temperature. The precipitate wasfiltered off and the solution was treated with silica and evaporatedunder reduced pressure and was purified by flash column chromatographyeluting with 100% cyclohexane gradually increasing to 30% EtOAc incyclohexane. Appropriate fractions were evaporated under reducedpressure to yield methyl2-bromo-2-[4-fluoro-3-(trifluoromethoxy)phenyl]acetate as a pale yellowoil (1.1 g, 129%).

¹H NMR (400 MHz, CDCl₃-d6, 25° C.) δ 3.81 (3H, s), 5.29 (1H, s),7.17-7.24 (1H, m), 7.46-7.52 (1H, m), 7.53-7.59 (1H, m).

Intermediate 55 Methyl 2-[4-fluoro-3-(trifluoromethoxy)phenyl]acetate

4-Fluoro-3-(trifluoromethoxy)phenylacetic acid (1.0 g, 4.199 mmol) wassuspended in methanol (10 mL) and treated with sulfuric acid (0.07 mL,0.84 mmol) and heated at 45° C. for 2 hours. The reaction mixture wasallowed to cool to room temperature and the methanol removed underreduced pressure. The residue was diluted with brine (20 mL) and thenextracted with EtOAc (3×20 mL). The combined EtOAc extracts were washedwith brine (30 mL) dried over MgSO₄, filtered and the solvent wasremoved under reduced pressure to yield methyl2-[4-fluoro-3-(trifluoromethoxy)phenyl]acetate, (0.65 g, 61%)

¹H NMR (400 MHz, DMSO-d6) δ 3.63 (3H, s), 3.78 (2H, s), 7.33-7.39 (1H,m), 7.42-7.53 (2H, m). m/z: ES⁺ [M+H]⁺ 253.

Biological Assays

The following assays were used to measure the effects of the compoundsdescribed herein: a) GLS Enzyme Potency Assay; b) GLS Cell PotencyAssay; c) GLS Cell Proliferation Assay. During the description of theassays, generally:

-   -   i. The following abbreviations have been used: CO₂=Carbon        dioxide; DMEM=Dulbecco's Modified Eagle Medium; DMSO=Dimethyl        sulphoxide; EDTA=Ethylenediaminetetraacetic acid; EGTA=Ethylene        glycol tetraacetic acid; FCS=Foetal calf serum; h=Hour(s);        NBS=Non-binding surface; SDS=Sodium dodecyl sulphate;        TRIS=Tris(Hydroxymethyl)aminomethane.    -   ii. IC₅₀ values were calculated using a smart fitting model in        Genedata. The IC₅₀ value was the concentration of test compound        that inhibited 50% of biological activity.

Assay a): GLS Enzyme Potency Assay

A Glutamate Oxidase/AmplexRed coupled assay was used to measure theability of compounds to bind to and inhibit the activity of GLS1 invitro. 6His tagged GLS protein (amino acids 63-669) expressed in E. Coliwas purified and stored at −80° C. in aliquots. GLS1 was diluted to 2×working concentration and incubated at room temperature to allow thetetrameric/dimeric forms to reach steady state. Assay measurements wereperformed in buffer comprising 50 mM TRIS pH 7.8, 100 mM NaPO₄, pH 7.8,0.001% v/v Tween20. Purified recombinant GLS1 protein was diluted inassay buffer to 12 nM and pre-incubated at room temperature for 30minutes. Test compounds were prepared by dilution in 100% DMSO to givethe correct dose range for 12 point concentration response and anappropriate volume (2.5-60 nl) dispensed into 384 well micro assayplates (Greiner product code 784900) using a Labcyte Echo 555 acousticdispenser. DMSO concentration was maintained at 2% by back filling withDMSO solution. 3 L of diluted GLS1 protein (12 nM) was then dispensedinto each well using a BioRaptr automated dispenser (Beckman-Coulter)and incubated for 15 minutes at room temperature. 3 L of 100 mMglutamine diluted in assay buffer was then added and the reactionincubated at room temperature for 60 minutes. The reaction was thenstopped by addition of 45 M6-(2-bromoethynyl)-2,3-dimethyl-quinazolin-4-one, 75 M Amplex Red, 0.375units/mL Horseradish Peroxidase, 0.12 units/mL Glutamate Oxidase in 100mM TRIS pH7.5. After 30 minutes at room temp in the dark, plates wereread on a Perkin Elmer EnVision using 535/590 nm optic filters and rawdata analysed using Genedata to generate IC₅₀ values. An artefactversion of the assay where the 6His tagged GLS protein and glutaminewere replaced with assay buffer was also used to rule out non specificeffects on the assay components.

Assay b): GLS Cell Potency Assay

Compounds were assessed for their potential to inhibit cellular GLSactivity by use of a PC3 coupled assay measuring cellular glutamatedepletion. Test compounds were prepared by dilution in 100% DMSO to givethe correct dose range for 12 point concentration response and anappropriate volume (5-120n) dispensed into 384 well micro assay plates(Corning product code 3712) using a Labcyte Echo 555 acoustic dispenser.DMSO concentration was maintained at 0.3% by back filling with DMSOsolution. PC3 cells were grown in phenol free DMEM, 10% dialyzed FCS, 2mM glutamine and following dispersal by trypsinisation were plated at5.6×10³ cells per well in 40 μl of growth medium directly into the 384well assay plates containing dispensed compound. After incubation for 6h at 37° C., 5% CO₂ growth media was aspirated and cells lysed in 15 μlof buffer containing 10 mM TRIS pH7.4, 100 mM NaCl, 1 mM EDTA, 1 mMEGTA, 1 mM NaF, 20 mM Na₄P₂O₇, 2 mM Na₃VO₄, 1% Triton X-100, 10%glycerol, 0.1% SDS and 0.5% deoxycholate. 4 μl of cell lysate was thentransferred to a 384 well NBS plate (Corning product code 3575) and 35μl of 27.5 μM Amplex Red, 0.1375 U/mL Horseradish Peroxidase, 0.044 U/mLglutamate oxidase, 100 mM TRIS pH7.5 was added. After 30 minutes at roomtemp in the dark, plates were read on a Perkin Elmer EnVision using535/590 nm optic filters and raw data analysed using proprietarysoftware to generate IC₅₀ values.

Assay c): GLS Cell Proliferation Assay

The ability of compounds to inhibit cell growth was measured using a 384well plate NCI-H1703 cell proliferation assay. NCI-H1703 cells weregrown in phenol red free RPMI1640, 10% FCS and 2 mM glutamine and seededat a density of 750 cells per well in 40 μl of growth medium intoclear-bottom 384 well assay plates (Corning product code 3712) andincubated for 24 h at 37° C., 5% CO₂. Test compounds were prepared bydilution in 100% DMSO to give the correct dose range for 12 pointconcentration response and an appropriate volume (5-120 nl) dispenseddirectly into the assay plates containing plated cells. DMSOconcentration was maintained at 0.3% by back filling with DMSO solution.Plates were incubated for 5 days at 37° C., 5% CO₂, Sytox Green andSaponin added to final concentration of 2 M and 0.25% respectively andincubated for 6 h prior to analysis. Plates were read on an Acumen eX3(TTP Labtech) using 488 nm excitation and FITC filter set (500-530 nm)for emission. IC₅₀ values were calculated by curve fitting to maxinhibition of day zero growth using GeneData software analysis.

Results from assays a)-c) are shown in Table 1.

TABLE 1 Assay data Assay b) Assay c) Assay a) GLS cell Prolif enz IC₅₀MOA Mean Mean IC₅₀ Example μM IC₅₀ μM μM  1(a) 0.0206 0.00083 0.00255 1(b) 0.0726 0.0184 0.133  2(a) 0.0872 0.000245 0.0025  2(b) 0.1210.00411 0.0286  3(a) 0.0308 0.000823 0.00265  3(b) 0.0721 0.014 0.272 4(a) 0.0423 0.000809 0.00295  4(b) 0.0596 0.00608 0.0273  5(a) 0.02880.00122 0.0117  5(b) 0.13 0.0181 0.12  6(a) 0.028 0.000659 0.0022  6(b)0.213 0.0226 0.0659  7(a) — — 0.00164  7(b) 0.192 0.0485 0.151  8(a)0.0649 0.000664 0.0576  8(b) 0.19 0.0326 0.0385  9(a) 0.0591 0.001020.0049  9(b) 0.519 0.0364 0.122 10(a) 0.0349 — — 10(b) 0.0722 0.007870.0434 11(a) 0.00635 0.000229 0.0017 11(b) 0.00611 0.000309 0.0010412(a) 0.0438 0.00193 0.0329 12(b) 0.971 0.0836 0.524 13(a) 0.07510.00273 0.0142 13(b) 1.42 0.0499 0.497 14(a) 0.0915 0.000872 0.0058914(b) 0.66 0.0146 0.156 15(a) 0.0776 0.00174 0.0116 15(b) 1.31 0.07570.376 16(a) 0.0252 0.000664 0.00349 16(b) 0.689 0.0384 — 17(a) 0.0504 —0.00156 17(b) 0.157 0.00711 0.0349 18(a) 0.0226 0.00213 0.0135 18(b)0.129 0.0546 0.699 19(a) 0.0401 0.000473 0.00261 19(b) 0.0961 0.003070.00759 20(a) 0.0693 0.00207 0.00657 20(b) 0.0641 0.000841 0.00265 21(a)0.02 0.000292 0.00224 21(b) 0.0482 0.00885 0.0591 22(a) 0.0685 — 0.0053522(b) 0.131 0.0106 0.178 23 0.0482 0.00885 0.0591 24 0.0526 0.003480.0291 25 1.84 0.0861 0.831 26 0.0235 0.000334 0.0042 27 0.149 0.00140.0135 28 0.195 0.0515 0.262 29 0.0511 0.000404 0.00196 30 1.37 0.003220.0377 31 0.0968 0.000573 0.00321 32 0.188 0.0146 0.0987 33 0.0685 —0.00535 34 0.131 0.0106 0.018 35 0.077 0.00121 0.00903 36 0.612 0.02180.394 37 0.147 0.000556 0.0211 38 0.19 — 0.303 39 0.13 0.000405 0.0059440 0.311 — 0.0825 41 0.0467 0.000468 0.00146 42 0.206 — 0.0815 43 0.020.000292 0.00224 44 0.0482 0.00885 0.0591 45(a) 1.60 0.147 2.27 45(b)0.146 0.0113 0.00922

1. A compound of Formula (I):

or a pharmaceutically acceptable salt thereof, wherein: Q is1,2,4-triazin-3-yl, pyridazin-3-yl, 6-methylpyridazin-3-yl, or6-fluoropyridazin-3-yl; R¹ is methoxy, trifluoromethoxy, oxetan-3-yl,3-fluoroazetidin-1-yl, 3-methoxyazetidin-1-yl, or3,3-difluoroazetidin-1-yl; R² is hydrogen or fluoro; R³ is hydrogen ormethoxy; and R⁴ is methoxy, ethoxy, or methoxymethyl; provided that whenR¹ is methoxy or trifluoromethoxy, then R³ is not hydrogen, and/or R⁴ ismethoxymethyl. 2-3. (canceled)
 4. The compound of claim 1, or apharmaceutically acceptable salt thereof, wherein R² is H.
 5. Thecompound of claim 1, or a pharmaceutically acceptable salt thereof,wherein R³ is methoxy.
 6. The compound of claim 1, or a pharmaceuticallyacceptable salt thereof, wherein R⁴ is methoxy.
 7. The compound of claim1, or a pharmaceutically acceptable salt thereof, wherein Q is1,2,4-triazin-3-yl, or pyridazin-3-yl. 8-9. (canceled)
 10. Apharmaceutical composition comprising the compound of claim 1, or apharmaceutically acceptable salt thereof, and at least onepharmaceutically acceptable diluent or carrier. 11-15. (canceled)